Alain Dautant | Centre National de la Recherche Scientifique / French National Centre for Scientific Research (original) (raw)

Egyptology by Alain Dautant

Program of the conference, 2019

Clergés et cultes thébains des Libyens aux Saïtes - Musée de Grenoble Un colloque international ... more Clergés et cultes thébains des Libyens aux Saïtes - Musée de Grenoble

Un colloque international organisé par le musée de Grenoble, le musée du Louvre, Sorbonne Université et la Société française d'égyptologie

Theban Cults and Priesthood from the Libyan to the Saïte Periods
An International Conference organized by the Musée de Grenoble, the Musée du Louvre, Sorbonne Université and the Société française d'égyptologie

In 1891, on the indication of Mohammed Ahmed Abd-el-Rassoul, Eugène Grébaut found the Bab el-Gasu... more In 1891, on the indication of Mohammed Ahmed Abd-el-Rassoul, Eugène Grébaut found the Bab el-Gasus cache at Deir el-Bahari, full of funerary equipments from the second part of the 21st Dynasty. The “liste A” of 153 coffins established in the tomb by Georges Daressy is the main source of information on these coffins available today. It must be added drawings from originals vignettes performed by Philippe Virey after the transfer of these coffins in the Gizeh museum. Furthermore, the unpublished shipping lists of coffins and shabtis of the lot n° 1 offered by the khedivial government to France (Ministère de l’Instruction publique et des Beaux-arts), written by Emile Brugsch, informs us about its content. This lot contained two double and three single coffins, three shabti boxes, 92 shabtis and a sheet of linen used for embalming. In his book on the 21st Dynasty coffins, Niwinski located two ensembles in Amiens and Clermont-Ferrand , while the localization of others was unknown. The annotations on the Brugsch’s list reveal the destination of the objects. Amiens, Boulogne-sur-Mer, Clermont-Ferrand, the Louvre and Marseille received a coffin. Otherwise, analysis of the Brugsch’s list and Virey’s drawings reveals that France received coffins from six different ensembles and not five as expected from Daressy’s list. Moreover, two shabti boxes and twelve shabtis were sent to Toulouse, fourteen shabtis to Lyon and the linen to Boulogne-sur-Mer. Here, the current locations of the coffins, the shabtis and shabti boxes are presented.

In his corpus of the 21st/22nd Dynasties yellow coffins published in 1988, Andrzej Niwinski ident... more In his corpus of the 21st/22nd Dynasties yellow coffins published in 1988, Andrzej Niwinski identified 458 coffins disseminated in the museums of twenty-seven countries, among these 44 were in France. We present an updated list with 43 more coffins identified in public and private French museums and collections. Most of these 87 coffins arrived in France in the nineteenth century through antique dealers (Perrot, Raynaud) and collectors (Bois-Aymé, Cailliaud, de Chassiron, Clot Bey, Déchelette, Durand, Godard, de Moncabrié, de Montulé, de Saint-Ferriol, Thédenat-Duvent, Tyszkiewicz). Beside these transfers, they were gifts of the Egyptian government. Among these, the most important were the coffins from the Bab el-Gasus cache offered to France by the Khedivial government in 1893.

The outer coffin of Djedmut, chantress of Amun, is a masterpiece of the Museo Gregoriano Egizio (... more The outer coffin of Djedmut, chantress of Amun, is a masterpiece of the Museo Gregoriano Egizio (Vatican). Her inner coffin was never clearly identified. The lid of the inner coffin of Djedmut with the same titles conserved at the Museum d’Histoire Naturelle (MHN) in La Rochelle (France) and likely the floorboard fragment of the inner coffin case with the same name at the Museo Civico Archeologico di Padova (Italy) could be the membra disjecta of the funerary equipment of Djedmut. The title, #nmm-#nsw-pA-Xrd, could be the name of Djedmut.

In the first two decades of the 19th century the collect of ancient Egyptian antiquities was boom... more In the first two decades of the 19th century the collect of ancient Egyptian antiquities was booming. There is no doubt that 21st dynasty collective tombs were uncovered at Western Thebes by Gurnawis in autumn 1820. At that time, Thédenat-Duvent, Lebolo for Drovetti, d’Athanasi for Salt, Cailliaud, d’Anastasy, de Castiglioni, Picchianti, and von Minutoli were in Luxor and bought various lots of burial equipments that they sell subsequently in European countries (Austria, England, France, Germany, Italy, Russia). Today, the materials are dispersed in numerous museums of England (Bolton, Cambridge, London, Oxford), France (Avignon, Avranches, Beaune, Besançon, Cannes, Dijon, Lille, Lyon, Marseille, Montpellier, Nantes, BNF and Louvre in Paris, Rennes, Rouen, Saint-Omer, Sens), Italy (Bologna, Florence, Naples), Russia (Moscow, St. Petersburg), and also in Berlin, Leiden, Oslo, Vienna, Zagreb and in private collections. Mummy covers, inner and outer coffins, linen jackets, books of the Dead and of Amdouat, canopic boxes and cofinettes, Osiris figures, shabti boxes, wooden and blue faience ushebtis belong to at least six families: (1) Khonsumes, Tentamon, Djedmutiufankh and Tayuheret, (2) Bakenmut and Mutemiya, (3) Sutymes and Penimen, (4) Seramun, and likely (5) Henuttauy and (6) Chauenhuy. Similarities in the types of materials and the owner titles reveal connection between these families.

All the coffins presented by the Khedival government of Egypt to France (Lot no. 1) and Russia (L... more All the coffins presented by the Khedival government of Egypt to France (Lot no. 1) and Russia (Lot no. 6) in 1893 couldn't be located or identified in 1988. Thanks to the packing slips for lots 1 and 6 prepared by Emile Brugsch at the Gizeh Museum, to unpublished drawings made by Philippe Virey between 1892 and 1893 in preparation for the publication of Bab el-Gusus by Grebaut (which never appeared), and the examination of the coffins in French and Russian museums and in the storerooms of the Egyptian Museum in Cairo, it is now possible to correctly identify and relocate all the
coffins from these two lots. A mixup in the coffins of lots 1 and 6 occurred in Gizeh, and one coffin from lot 6 remained in the Gizeh Museum. Mikola Tarasenko (Kiev, Ukraine) and Andrzej Niwiflski (Warsaw University, Poland) collaborated in this research. Further
research has uncovered a unique photograph by Abdullah Freres taken in the Gizeh Museum during the unwrapping a mummy from Bab el-Gusus, showing Mashetseketeb's coffin lid placed on the floor beside it. Four other coffins of the 21st-22nd Dynasties (not from Bab el-Gusus) exhibited in the Gizeh and Cairo Museums have also been identified in old photographs.

South of Kom el-Samak, a second platform of mud brick, Kom el-Abd, rises out of the low desert. ... more South of Kom el-Samak, a second platform of mud brick, Kom el-Abd, rises out of the low desert. But this structure was not the end of Amenhotep’s plans for his the area. At some point near the end of his reign (it appears that he never completed the project) Amenhotep III cleared a giant strip in the low desert. The strip, identified by the large stones piled along its edges, extends to the foot of the terraces leading up to the cliffs of the high desert, the Sahara. The purpose of this strip is as yet undetermined.

First Vatican Coffin Conference, Jul 2013

In his corpus of the 21st Dynasty coffins published in 1988, Andrzej Niwinski identified 458 buri... more In his corpus of the 21st Dynasty coffins published in 1988, Andrzej Niwinski identified 458 burial sets disseminated in the museums of twenty-seven countries. A set can range from a double coffin with mummy-board to a single element (lid, case, board or fragment). A systematic and detailed study of Egyptian coffins of the Third Intermediate Period preserved in the French museums is underway. At least thirty-eight coffins (21st-22th Dynasty) arrived in France in the nineteenth century through antique dealers and collectors and five in 1893 from the second cachette of Deir el-Bahari.
We will present an updated list with a few recently published and unpublished coffins such as the Itneferamun coffin (Musée d'Aquitaine, Bordeaux) and the lid of the internal stola-type coffin of Djedmut (Museum Histoire Naturelle, La Rochelle) which certainly belongs to one of the masterpieces of the Gregorian Egyptian Museum, Vatican.

First Vatican Coffin Conference, Jul 2013

Scientific secretary Ma rio C appozzo, ~gypt i an Dept. Musei Vat ican i coffinconfe rence.musei@... more Scientific secretary Ma rio C appozzo, ~gypt i an Dept. Musei Vat ican i coffinconfe rence.musei@scv.va Organization ~vent Dept. and Web-Dept . M usei Vatican i Progetto grafico e impaginazione G iulia A ngeli ni info@ angelin ico muni caz io ne.it Peferenze fo tografiche f= ot o © M usei Vat icani Stampa Tip ografia Vatica na © Gove rnato riato SCV -Direz ion e d ei M use i Vat ican i C ittà d el Vatica no www. mu se iva ti ca ni .va TI-IANKS TO Massachusetts Chapter Patrons of the Arts in the Vatican Museums

Cercueils jaunes des XXIe et XXIIe dynasties dans les collections Françaises, 2014

In 1988, Andrzej Niwiski counted 44 Egyptian yellow coffins or fragments dating 21st and 22nd dyn... more In 1988, Andrzej Niwiski counted 44 Egyptian yellow coffins or fragments dating 21st and 22nd dynasties in French collections. Then, in 1997, René van Walsem added to the inventory four new coffins of stola type, i.e. coffins with red mummy braces crossing on the chest. In the present corpus, the total number was increased to 87; two of these coffins were abroad in 1988. In addition, six non-localized coffins (Niw 208, 427, 428, 450, 455 and 457) were identified, two coffins were identified (Niw 331 and 332) and the five sets from Bab el-Gasus offered to France in 1893 are located. This study therefore provides an updated list of 21st and 22nd dynasties yellow coffins in French collections and aims at clarifying the provenance of some coffins.

14.30 - Opening 15.00 - René van Walsem University of Leiden) - "MastaBase, a database for the... more 14.30 - Opening

15.00 - René van Walsem University of Leiden) - "MastaBase, a database for the quantitative analysis of Old Kingdom iconographic programmes and their texts in the elite tombs of the Memphite área".

15.30 - Kathlyn Cooney (University of California - Los Angeles)- "The Coffin Database Project".

16.00 - Christian Greco (Museum of Antiquities of Leiden) - "The *qrsw* coffins as cosmogram. Correspondence between the burial chamber of Ramose (TT 132) and the coffin of Ankhefenkhonsu".

16.30 - Coffee-break

17.00 - Rogério Sousa (CITCEM, University of Porto) - "Typology of coffin decoration in the 21st Dynasty: the contribution of the study of the Eight Lot of Bab el-Gasus".

17.30 - Alessia Amenta (Vatican Museums), Christian Greco (Leiden Museum of Antiquities), Helène Guichard (Louvre Museum) - "The Vatican Coffin Project: Goals and preliminary results".

Revue Archéologique de Bordeaux t.103, p.11-39, 2012

Résumé La collection d’antiquités égyptiennes du musée d’Aquitaine comporte près de 700 objets. ... more Résumé
La collection d’antiquités égyptiennes du musée d’Aquitaine comporte près de 700 objets. Elle s’est constituée au cours des XIXe et XXe siècle grâce à une succession de dons, legs et achats. L’histoire de cette collection se confond avec celle des musées de Bordeaux. Le cercueil thébain d’Irethorrou (XXVe dynastie), offert à la Société Archéologique de Bordeaux par C.A. Ducatel en 1877, fait l’objet de la présente étude.
Abstract
The collection of Egyptian antiquities of the Aquitaine museum contains about 700 objects. It was constituted during XIXth and XXth century thanks to a succession of donations, legacies and purchases. The history of this collection becomes confused with that of the Bordeaux’s museums. The Theban coffin of Irethorru (XXVth dynasty), offered to the Archaeological Society of Bordeaux by C.A. Ducatel in 1877, is presently studied.

Three-dimensional terahertz computed tomography has been used to investigate dried human bones su... more Three-dimensional terahertz computed tomography has been used to investigate dried human bones such as a lumbar vertebra, a coxal bone, and a skull, with a direct comparison with standard radiography. In spite of lower spatial resolution compared with x-ray, terahertz imaging clearly discerns a compact bone from a spongy one, with strong terahertz absorption as shown by additional terahertz time-domain transmission spectroscopy.

Program of the conference, 2019

Clergés et cultes thébains des Libyens aux Saïtes - Musée de Grenoble Un colloque international ... more Clergés et cultes thébains des Libyens aux Saïtes - Musée de Grenoble

Un colloque international organisé par le musée de Grenoble, le musée du Louvre, Sorbonne Université et la Société française d'égyptologie

First Vatican Coffin Conference, Jul 2013

Cercueils jaunes des XXIe et XXIIe dynasties dans les collections Françaises, 2014

14.30 - Opening 15.00 - René van Walsem University of Leiden) - "MastaBase, a database for the... more 14.30 - Opening

15.00 - René van Walsem University of Leiden) - "MastaBase, a database for the quantitative analysis of Old Kingdom iconographic programmes and their texts in the elite tombs of the Memphite área".

15.30 - Kathlyn Cooney (University of California - Los Angeles)- "The Coffin Database Project".

16.00 - Christian Greco (Museum of Antiquities of Leiden) - "The *qrsw* coffins as cosmogram. Correspondence between the burial chamber of Ramose (TT 132) and the coffin of Ankhefenkhonsu".

16.30 - Coffee-break

17.00 - Rogério Sousa (CITCEM, University of Porto) - "Typology of coffin decoration in the 21st Dynasty: the contribution of the study of the Eight Lot of Bab el-Gasus".

17.30 - Alessia Amenta (Vatican Museums), Christian Greco (Leiden Museum of Antiquities), Helène Guichard (Louvre Museum) - "The Vatican Coffin Project: Goals and preliminary results".

Revue Archéologique de Bordeaux t.103, p.11-39, 2012

Int J Mol Sci, 2020

Nucleoside diphosphate kinases (NDPK) are oligomeric proteins involved in the synthesis of nucleo... more Nucleoside diphosphate kinases (NDPK) are oligomeric proteins involved in the synthesis of nucleoside triphosphates. Their tridimensional structure has been solved by X-ray crystallography and shows that individual subunits present a conserved ferredoxin fold of about 140 residues in prokaryotes, archaea, eukaryotes and viruses. Monomers are functionally independent from each other inside NDPK complexes and the nucleoside kinase catalytic mechanism involves transient phosphorylation of the conserved catalytic histidine. To be active, monomers must assemble into conserved head to tail dimers, which further assemble into hexamers or tetramers. The interfaces between these oligomeric states are very different but, surprisingly, the assembly structure barely affects the catalytic efficiency of the enzyme. While it has been shown that assembly into hexamers induces full formation of the catalytic site and stabilizes the complex, it is unclear why assembly into tetramers is required for function. Several additional activities have been revealed for NDPK, especially in metastasis spreading, cytoskeleton dynamics, DNA binding and membrane remodeling. However, we still lack the high resolution structural data of NDPK in complex with different partners, which is necessary for deciphering the mechanism of these diverse functions. In this review we discuss advances in the structure, folding and stability of NDPKs.

Biochemistry, 2019

In order to be fully active and participate in the metabolism of phosphorylated nucleotides, most... more In order to be fully active and participate in the metabolism of phosphorylated nucleotides, most nucleoside diphosphate kinases (NDPK) have to assemble into stable hexamers. Here we studied the role played by six inter-subunit salt bridges R80-D93 in the stability of NDPK from the pathogen Mycobacterium tuberculosis ( Mt). Mutating R80 into Ala or Asn abolished the salt bridges. Unexpectedly, compensatory stabilizing mechanisms appeared for R80A and R80N mutants and we studied them by biochemical and structural methods. R80A mutant crystallized into I222 space group unusual for NDPK and its hexameric structure revealed occurrence at the trimer interface of a stabilizing hydrophobic patch around the mutation. Functionally relevant, a trimer of the R80A hexamer showed a remodeling of the binding site. In this conformation, the cleft of active site is more open, and then active His117 is more accessible to substrates. HDX-MS analysis of WT, R80A and R80N mutants showed that the remodeled region of the protein is highly solvent accessible indicating that equilibrium between open and closed conformation is possible. We propose that such equilibrium occurs in vivo and explains how bulky substrates access the catalytic His117.

Biochemistry, 2017

Most oligomeric proteins become active only after assembly, but why oligomerization is required t... more Most oligomeric proteins become active only after assembly, but why oligomerization is required to support function is not well understood. Here, we address this question using the wild type (WT) and a destabilized mutant (D93N) of the hexameric nucleoside diphosphate kinase from the pathogen Mycobacterium tuberculosis (Mt-NDPK). The conformational dynamics and oligomeric states of each were analyzed during unfolding and/or folding by hydrogen/deuterium exchange mass spectrometry (HDX-MS) at peptide resolution and by additional biochemical techniques. We found that WT and D93N native hexamers present a stable core and a flexible periphery, the latter being more flexible for the destabilized mutant. Stable but inactive species formed during unfolding of D93N and folding of WT were characterized. For the first time, we show that both of these species are nativelike dimers, each of its monomers having a major subdomain folded, while a minor subdomain (Kpn/α0) remains unfolded. The Kpn/α0 subdomain, which belongs to the catalytic site, becomes structured only upon hexamerization, explaining why oligomerization is required for NDPK activity. Further HDX-MS studies are necessary to establish the general activation mechanism for other homo-oligomers.

The crystal structure of the wild-type nucleoside diphosphate kinase from Mycobacterium tuberculo... more The crystal structure of the wild-type nucleoside diphosphate kinase from Mycobacterium tuberculosis at 2.6 Ang resolution revealed that the intersubunit salt bridge Arg80–Asp93 contributes to the thermal stability of the hexamer (Tm = 76°C). On mutating Asp93 to Asn to break the salt bridge, the thermal stability dramatically decreased by 27.6°C. Here, on mutating Arg80 to Asn, the thermal stability also significantly decreased by 8.0°C. In the X-ray structure of the R80N mutant solved at 1.9 Ang resolution the salt bridge was replaced by intersubunit hydrogen bonds that contribute to the thermal stability of the hexamer. A citrate anion from the crystallization buffer was bound at the bottom of the nucleotide-binding site via electrostatic and hydrogen-bonding interactions with six conserved residues involved in nucleotide binding. Structural analysis shows that the citrate is present at the location of the nucleotide phosphate groups.

Abstract Most nucleoside diphosphate kinases (NDPKs) are hexamers. The C-terminal tail interactin... more Abstract Most nucleoside diphosphate kinases (NDPKs) are hexamers. The C-terminal tail interacting with the neighboring subunits is crucial for hexamer stability. In the NDPK from Mycobacterium tuberculosis (Mt) this tail is missing. The quaternary structure of Mt-NDPK is essential for full enzymatic activity and for protein stability to thermal and chemical denaturation. We identified the intersubunit salt bridge Arg 80-Asp 93 as essential for hexamer stability, compensating for the decreased intersubunit contact area.

The nucleoside diphosphate kinase (Ndk) catalyzes the reversible transfer of the γ-phosphate from... more The nucleoside diphosphate kinase (Ndk) catalyzes the reversible transfer of the γ-phosphate from nucleoside triphosphate to nucleoside diphosphate. Ndks form hexamers or two types of tetramers made of the same building block, namely, the common dimer. The secondary interfaces of the Type I tetramer found in Myxococcus xanthus Ndk and of the Type II found in Escherichia coli Ndk involve the opposite sides of subunits. Up to now, the few available structures of Ndk from thermophiles were hexameric. Here, we determined the X-ray structures of four crystal forms of the Ndk from the hyperthermophilic bacterium Aquifex aeolicus (Aa-Ndk). Aa-Ndk displays numerous features of thermostable proteins and is made of the common dimer but it is a tetramer of Type I. Indeed, the insertion of three residues in a surface-exposed spiral loop, named the Kpn-loop, leads to the formation of a two-turn α-helix that prevents both hexamer and Type II tetramer assembly. Moreover, the side chain of the cysteine at position 133, which is not present in other Ndk sequences, adopts two alternate conformations. Through the secondary interface, each one forms a disulfide bridge with the equivalent Cys133 from the neighboring subunit. This disulfide bridge was progressively broken during X-ray data collection by radiation damage. Such crosslinks counterbalance the weakness of the common-dimer interface. A 40% decrease of the kinase activity at 60°C after reduction and alkylation of the protein corroborates the structural relevance of the disulfide bridge on the tetramer assembly and enzymatic function.

Journal of Structural …, Jan 1, 2011

The F(1)F(O)-ATP synthase is a rotary molecular nanomotor. F(1) is a chemical motor driven by ATP... more The F(1)F(O)-ATP synthase is a rotary molecular nanomotor. F(1) is a chemical motor driven by ATP hydrolysis while F(O) is an electrical motor driven by the proton flow. The two stepping motors are mechanically coupled through a common rotary shaft. Up to now, the three available crystal structures of the F(1)c(10) sub-complex of the yeast F(1)F(O)-ATP synthase were isomorphous and then named yF(1)c(10)(I). In this crystal form, significant interactions of the c(10)-ring with the F(1)-head of neighboring molecules affected the overall conformation of the F(1)-c-ring complex. The symmetry axis of the F(1)-head and the inertia axis of the c-ring were tilted near the interface between the F(1)-central stalk and the c-ring rotor, resulting in an unbalanced machine. We have solved a new crystal form of the F(1)c(10) complex, named yF(1)c(10)(II), inhibited by adenylyl-imidodiphosphate (AMP-PNP) and dicyclohexylcarbodiimide (DCCD), at 6.5Å resolution in which the crystal packing has a weaker influence over the conformation of the F(1)-c-ring complex. yF(1)c(10)(II) provides a model of a more efficient generator. yF(1)c(10)(II) and bovine bF(1)c(8) structures share a common rotor architecture with the inertia center of the F(1)-stator close to the rotor axis.

Hypoxanthine-guanine phosphoribosyltransferase (HGPRT) is a key enzyme of the purine recycling pa... more Hypoxanthine-guanine phosphoribosyltransferase (HGPRT) is a key enzyme of the purine recycling pathway that catalyzes the conversion of 5-phospho-ribosyl-α-1-pyrophosphate and guanine or hypoxanthine to guanosine monophosphate (GMP) or inosine monophosphate (IMP), respectively, and pyrophosphate (PPi). We report the first crystal structure of a fungal 6-oxopurine phosphoribosyltransferase, the Saccharomyces cerevisiae HGPRT (Sc-HGPRT) in complex with GMP. The crystal structures of full length protein with (WT1) or without (WT2) sulfate that mimics the phosphate group in the PPi binding site were solved by molecular replacement using the structure of a truncated version (Δ7) solved beforehand by multiwavelength anomalous diffusion. Sc-HGPRT is a dimer and adopts the overall structure of class I phosphoribosyltransferases (PRTs) with a smaller hood domain and a short two-stranded parallel β-sheet linking the N- to the C-terminal end. The catalytic loops in WT1 and WT2 are in an open form while in Δ7, due to an inter-subunit disulfide bridge, the catalytic loop is in either an open or closed form. The closure is concomitant with a peptide plane flipping in the PPi binding loop. Moreover, owing the flexibility of a GGGG motif conserved in fungi, all the peptide bonds of the phosphate binding loop are in trans conformation whereas in nonfungal 6-oxopurine PRTs, one cis-peptide bond is required for phosphate binding. Mutations affecting the enzyme activity or the previously characterized feedback inhibition by GMP are located at the nucleotide binding site and the dimer interface.

C262 docking techniques and molecular dynamics simulations. Modeling results indicate that positi... more C262 docking techniques and molecular dynamics simulations. Modeling results indicate that positions C7 and C10 affect the conformation of three key elements (the M-loop, S3 and H3) in the lateral interactions that modulate the contacts between adjacent protofilaments. Alternatively, the change in C2 slightly rearranges the ligand in the binding site, thus modifying the interaction of the ligand C7 position with the M-loop.

Journal of Biological Chemistry, Jan 1, 2010

The F(1)c(10) subcomplex of the yeast F(1)F(0)-ATP synthase includes the membrane rotor part c(10... more The F(1)c(10) subcomplex of the yeast F(1)F(0)-ATP synthase includes the membrane rotor part c(10)-ring linked to a catalytic head, (αβ)(3), by a central stalk, γδε. The Saccharomyces cerevisiae yF(1)c(10)·ADP subcomplex was crystallized in the presence of Mg·ADP, dicyclohexylcarbodiimide (DCCD), and azide. The structure was solved by molecular replacement using a high resolution model of the yeast F(1) and a bacterial c-ring model with 10 copies of the c-subunit. The structure refined to 3.43-Å resolution displays new features compared with the original yF(1)c(10) and with the yF(1) inhibited by adenylyl imidodiphosphate (AMP-PNP) (yF(1)(I-III)). An ADP molecule was bound in both β(DP) and β(TP) catalytic sites. The α(DP)-β(DP) pair is slightly open and resembles the novel conformation identified in yF(1), whereas the α(TP)-β(TP) pair is very closed and resembles more a DP pair. yF(1)c(10)·ADP provides a model of a new Mg·ADP-inhibited state of the yeast F(1). As for the original yF(1) and yF(1)c(10) structures, the foot of the central stalk is rotated by ∼40 ° with respect to bovine structures. The assembly of the F(1) central stalk with the F(0) c-ring rotor is mainly provided by electrostatic interactions. On the rotor ring, the essential cGlu(59) carboxylate group is surrounded by hydrophobic residues and is not involved in hydrogen bonding.

Pour vivre, nous avons besoin d'adénosine 5’-triphosphate, plus connu sous l’acronyme ATP. Mais c... more Pour vivre, nous avons besoin d'adénosine 5’-triphosphate, plus connu sous l’acronyme ATP. Mais comment cette molécule est-elle synthétisée ? C'est à cette question qu'essaye de répondre une équipe de l'Institut de Biologie et Génétique Cellulaires (CNRS/Université Bordeaux 2) en étudiant, à l'échelle atomique et moléculaire, la H+-F1F0 ATP synthase mitochondriale de la levure Saccharomyces cerevisiae. Les chercheurs ont réussi à cristalliser et résoudre la structure cristalline d'un sous-complexe de cette enzyme. Leurs résultats ont été publiés le 17 septembre 2010 dans The Journal of Biological Chemistry.

The humanin family of peptides have demonstrated in vivo and in vitro neuroprotective action agai... more The humanin family of peptides have demonstrated in vivo and in vitro neuroprotective action against the insults of Alzheimer's disease, offering thus a new perspective for the development of pharmaceutical agents against Alzheimer's disease. In this work, three members of the family, the parent peptide humanin (HN, MAPRG-FSCLLLLTSEIDLPVKRRA), its potent analogue HNG having a Gly in the place of Ser14, as well as a hybrid derivative (Colivelin, CL, SALLRSIPAPAGASRLLLLTGEIDLP) produced by the combination of the activity dependent neurotrophic factor (SALLRSIPA) and the transformed active core of HN (PAGASRLLLLTGEIDLP), are investigated for their solution structure, their in vitro affinities for neuronal cell lines and brain tissue and their in vivo biodistribution. The solution structure is approached with NMR and CD methodologies while the in vivo and in vitro biological experiments are performed through the use of 125 I-radiolabelled derivatives. The structural results reveal flexible peptides in the aqueous environment able -under certain conditions -to self-associate through the formation of b-sheet structures, a property that is correlated in the literature with their neuroprotective potency. The biological evaluation studies reveal much higher stability in serum than in tissue homogenates, lack of any obvious specific binding in the brain tissue, and a significant accumulation in the stomach, a fact that is currently further investigated. These findings will be discussed and compared to the properties of other agents of related action in the effort to elucidate the important structural and biochemical features that determine the neuroprotective action of this family of peptides.

Journal of bioenergetics …, Jan 1, 2006

Nm23 was the first metastasis suppressor gene identified. This gene encodes a NDP kinase that als... more Nm23 was the first metastasis suppressor gene identified. This gene encodes a NDP kinase that also exhibits other properties like histidine protein kinase and interactions with proteins and DNA. The S120G mutant of NDPK-A has been identified in aggressive neuroblastomas and has been found to reduce the metastasis suppressor effect of Nm23. In order to understand the differences between the wild type and the S120G mutant, we have determined the structure of both mutant and wild type NDPK-A in complex with ADP. Our results reveal that there are no significant changes between the two enzyme versions even in the surroundings of the catalytic histidine that is required for NDP kinase activity. This suggests that the S120G mutation may affect an other protein property than NDP kinase activity.

Proteins: Structure, …, Jan 1, 2007

Nucleoside diphosphate kinase (NDPK) catalyzes the transfer of gamma-phosphate from nucleoside tr... more Nucleoside diphosphate kinase (NDPK) catalyzes the transfer of gamma-phosphate from nucleoside triphosphates to nucleoside diphosphates. The subunit folding and the dimeric basic structural unit are remarkably the same for available structures but, depending on species, dimers self-associate to form hexamers or tetramers. The crystal structure of the Escherichia coli NDPK reveals a new tetrameric quaternary structure for this protein family. The two tetramers differ by the relative orientation of interacting dimers, which face either the convex or the concave side of their central sheet as in either Myxococcus xanthus (type I) or E. coli (type II), respectively. In the type II tetramer, the subunits interact by a new interface harboring a zone called the Kpn loop as in hexamers, but by the opposite face of this loop. The evolutionary conservation of the interface residues indicates that this new quaternary structure seems to be the most frequent assembly mode in bacterial tetrameric NDP kinases.

… Section D: Biological …, Jan 1, 2001

Cubic F432 crystals of recombinant mouse L-chain apoferritin were obtained by the hanging-drop te... more Cubic F432 crystals of recombinant mouse L-chain apoferritin were obtained by the hanging-drop technique with ammonium sulfate and cadmium sulfate as precipitants. The structure was refined to 2.1 and 1.6 A resolution from data obtained at room temperature and under cryogenic conditions, respectively. The structure of an eight-amino-acid loop insertion in the mouse sequence is found to be highly disordered both at room temperature and at low temperature.

The synthesis and X-ray structure analysis of a new manganese(III) complex [MnL1(H2O)2]OTf displa... more The synthesis and X-ray structure analysis of a new manganese(III) complex [MnL1(H2O)2]OTf displaying structural analogies with metalloporphyrins are described. This compound is characterized by very short Mn–N bond distances (1.93 Å) and a nearly planar structure. Its ability to catalyze cyclohexene epoxidation in the presence of MCPBA has also been evaluated.

European Journal …, Jan 1, 2000

Doxorubicin is among the most widely used anthracycline in cancer chemotherapy. In an attempt to ... more Doxorubicin is among the most widely used anthracycline in cancer chemotherapy. In an attempt to avoid the cardiotoxicity and drug resistance of doxorubicin therapy, several analogues were synthesized. The cyanomorpholinyl derivative is the most cytotoxic. They differ greatly from their parent compound in their biological and pharmacological properties, inducing cross-links in drug DNA complexes. The present study concerns N-cyanomethyl-N-(2-methoxyethyl)-daunomycin (CMDa), a synthetic analogue of cyanomorpholino-daunomycin. Compared to doxorubicin, CMDa displays a cytotoxic activity on L1210 leukemia cells at higher concentration but is effective on doxorubicin resistant cells. The results of fluorescence quenching experiments as well as the melting temperature (DeltaTm = 7.5 degrees C) studies are consistent with a drug molecule which intercalates between the DNA base pairs and stabilizes the DNA double helix. The crystal structure of CMDa complexed to the hexanucleotide d(CGATCG) has been determined at 1.5 A resolution. The complex crystallizes in the space group P41212 and is similar to other anthracycline-hexanucleotide complexes. In the crystal state, the observed densities indicate the formation of N-hydroxymethyl-N-(2-methoxyethyl)-daunomycin (HMDa) with the release of the cyano moiety without DNA alkylation. The formation of this degradation compound is discussed in relation with other drug modifications when binding to DNA. Comparison with two other drug-DNA crystal structures suggests a correlation between a slight change in DNA conformation and the nature of the amino sugar substituents at the N3' position located in the minor groove.

$Research paper thumbnail of Granier et al (2000) Acta Cryst D56 - Crystallization and preliminary X-ray diffraction data of mouse L-chain apoferritin crystals$

… Section D: Biological …, Jan 1, 2000

Crystals of recombinant mouse L-chain apoferritin were obtained by the hanging-drop technique usi... more Crystals of recombinant mouse L-chain apoferritin were obtained by the hanging-drop technique using ammonium sulfate as precipitant. Two crystal forms were observed in the same drop. The crystals belong to either the P2 monoclinic or to the P42(1)2 tetragonal space group. The monoclinic crystals diffracted to beyond 2.4 A resolution but were systematically twinned, while the tetragonal crystals diffracted to beyond 2.9 A. These crystallization conditions in the absence of metal salts should facilitate the study of the interaction between L-chain ferritins and heavy metals, particularly the iron core.

Human Molecular Genetics, 2021

The human ATP synthase is an assembly of 29 subunits of 18 different types, of which only two (a ... more The human ATP synthase is an assembly of 29 subunits of 18 different types, of which only two (a and 8) are encoded in the mitochondrial genome. Subunit a, together with an oligomeric ring of c-subunit (c-ring), forms the proton pathway responsible for the transport of protons through the mitochondrial inner membrane, coupled to rotation of the c-ring and ATP synthesis. Neuromuscular diseases have been associated to a number of mutations in the gene encoding subunit a, ATP6. The most common, m.8993 T > G, leads to replacement of a strictly conserved leucine residue with arginine (aL 156 R). We previously showed that the equivalent mutation (aL 173 R) dramatically compromises respiratory growth of Saccharomyces cerevisiae and causes a 90% drop in the rate of mitochondrial ATP synthesis. Here, we isolated revertants from the aL 173 R strain that show improved respiratory growth. Four first-site reversions at codon 173 (aL 173 M, aL 173 S, aL 173 K and aL 173 W) and five second-site reversions at another codon (aR 169 M, aR 169 S, aA 170 P, aA 170 G and aI 216 S) were identified. Based on the atomic structures of yeast ATP synthase and the biochemical properties of the revertant strains, we propose that the aL 173 R mutation is responsible for unfavorable electrostatic interactions that prevent the release of protons from the c-ring into a channel from which protons move from the c-ring to the mitochondrial matrix. The results provide further evidence that yeast aL 173 (and thus human aL 156) optimizes the exit of protons from ATP synthase, but is not essential despite its strict evolutionary conservation.

Life (Basel), 2020

With the advent of next generation sequencing, the list of mitochondrial DNA (mtDNA) mutations id... more With the advent of next generation sequencing, the list of mitochondrial DNA (mtDNA) mutations identified in patients rapidly and continuously expands. They are frequently found in a limited number of cases, sometimes a single individual (as with the case herein reported) and in heterogeneous genetic backgrounds (heteroplasmy), which makes it difficult to conclude about their pathogenicity and functional consequences. As an organism amenable to mitochondrial DNA manipulation, able to survive by fermentation to loss-of-function mtDNA mutations, and where heteroplasmy is unstable, Saccharomyces cerevisiae is an excellent model for investigating novel human mtDNA variants, in isolation and in a controlled genetic context. We herein report the identification of a novel variant in mitochondrial ATP6 gene, m.8909T>C. It was found in combination with the well-known pathogenic m.3243A>G mutation in mt-tRNALeu. We show that an equivalent of the m.8909T>C mutation compromises yeast adenosine tri-phosphate (ATP) synthase assembly/stability and reduces the rate of mitochondrial ATP synthesis by 20-30% compared to wild type yeast. Other previously reported ATP6 mutations with a well-established pathogenicity (like m.8993T>C and m.9176T>C) were shown to have similar effects on yeast ATP synthase. It can be inferred that alone the m.8909T>C variant has the potential to compromise human health.

Int J Mol Sci, 2020

Probing the pathogenicity and functional consequences of mitochondrial DNA (mtDNA) mutations from... more Probing the pathogenicity and functional consequences of mitochondrial DNA (mtDNA) mutations from patient's cells and tissues is difficult due to genetic heteroplasmy (co-existence of wild type and mutated mtDNA in cells), occurrence of numerous mtDNA polymorphisms, and absence of methods for genetically transforming human mitochondria. Owing to its good fermenting capacity that enables survival to loss-of-function mtDNA mutations, its amenability to mitochondrial genome manipulation, and lack of heteroplasmy, Saccharomyces cerevisiae is an excellent model for studying and resolving the molecular bases of human diseases linked to mtDNA in a controlled genetic background. Using this model, we previously showed that a pathogenic mutation in mitochondrial ATP6 gene (m.9191T>C), that converts a highly conserved leucine residue into proline in human ATP synthase subunit a (aL222P), severely compromises the assembly of yeast ATP synthase and reduces by 90% the rate of mitochondrial ATP synthesis. Herein, we report the isolation of intragenic suppressors of this mutation. In light of recently described high resolution structures of ATP synthase, the results indicate that the m.9191T>C mutation disrupts a four α-helix bundle in subunit a and that the leucine residue it targets indirectly optimizes proton conduction through the membrane domain of ATP synthase.

Int J Mol Sci, 2019

Bcl-x L is an oncogene of which the survival functions are finely tuned by post-translational mod... more Bcl-x L is an oncogene of which the survival functions are finely tuned by post-translational modifications (PTM). Within the Bcl-2 family of proteins, Bcl-x L shows unique eligibility to deamidation, a time-related spontaneous reaction. Deamidation is still a largely overlooked PTM due to a lack of easy techniques to monitor Asn→Asp/IsoAsp conversions or Glu→Gln conversions. Being able to detect PTMs is essential to achieve a comprehensive description of all the regulatory mechanisms and functions a protein can carry out. Here, we report a gel composition improving the electrophoretic separation of deamidated forms of Bcl-x L generated either by mutagenesis or by alkaline treatment. Importantly, this new gel formulation proved efficient to provide the long-sought evidence that even doubly-deamidated Bcl-x L remains eligible for regulation by phosphorylation.

Biochim Biophys Acta Bioenerg, 2019

Dozens of pathogenic mutations have been localized in the mitochondrial gene (MT-ATP6) that encod... more Dozens of pathogenic mutations have been localized in the mitochondrial gene (MT-ATP6) that encodes the subunit a of ATP synthase. The subunit a together with a ring of identical subunits c moves protons across the mitochondrial inner membrane coupled to rotation of the subunit c-ring and ATP synthesis. One of these mutations, m.8851T>C, has been associated with bilateral striatal lesions of childhood (BSLC), a group of rare neurological disorders characterized by symmetric degeneration of the corpus striatum. It converts a highly conserved tryptophan residue into arginine at position 109 of subunit a (aW109R). We previously showed that an equivalent thereof in Saccharomyces cerevisiae (aW126R) severely impairs by an unknown mechanism the functioning of ATP synthase without any visible assembly/stability defect. Herein we show that ATP synthase function was recovered to varying degree by replacing the mutant arginine residue 126 with methionine, lysine or glycine or by replacing with methionine an arginine residue present in position 169 of subunit a (aR169). In recently described atomic structures of yeast ATP synthase, aR169 is at the center of an hydrophilic cleft along which protons are transported from the subunit c-ring to the mitochondrial matrix, in the proximity of the two residues known from a long time to be essential to the activity of FO (aR176 and cE59). We provide evidence that the aW126R change is responsible for an electrostatic and steric hindrance that enables aR169 to engage in a salt bridge with cE59. As a result, aR176 cannot interact properly with cE59 and ATP synthase fails to effectively move protons across the mitochondrial membrane. In addition to insight into the pathogenic mechanism induced by the m.8851T>C mutation, the present study brings interesting information on the role of specific residues of subunit a in the energy-transducing activity of ATP synthase.

Biochim Biophys Acta Bioenerg, 2018

The ATP synthase which provides aerobic eukaryotes with ATP, organizes into a membrane-extrinsic ... more The ATP synthase which provides aerobic eukaryotes with ATP, organizes into a membrane-extrinsic catalytic domain, where ATP is generated, and a membrane-embedded FO domain that shuttles protons across the membrane. We previously identified a mutation in the mitochondrial MT-ATP6 gene (m.8969G>A) in a 14-year-old Chinese female who developed an isolated nephropathy followed by brain and muscle problems. This mutation replaces a highly conserved serine residue into asparagine at amino acid position 148 of the membrane-embedded subunit a of ATP synthase. We showed that an equivalent of this mutation in yeast (aS175N) prevents FO-mediated proton translocation. Herein we identified four first-site intragenic suppressors (aN175D, aN175K, aN175I, and aN175T), which, in light of a recently published atomic structure of yeast FO indicates that the detrimental consequences of the original mutation result from the establishment of hydrogen bonds between aN175 and a nearby glutamate residue (aE172) that was proposed to be critical for the exit of protons from the ATP synthase towards the mitochondrial matrix. Interestingly also, we found that the aS175N mutation can be suppressed by second-site suppressors (aP12S, aI171F, aI171N, aI239F, and aI200M), of which some are very distantly located (by 20-30 Å) from the original mutation. The possibility to compensate through long-range effects the aS175N mutation is an interesting observation that holds promise for the development of therapeutic molecules.

Biochim Biophys Acta Bioenerg, 2019

Protons are transported from the mitochondrial matrix to the intermembrane space of mitochondria ... more Protons are transported from the mitochondrial matrix to the intermembrane space of mitochondria during the transfer of electrons to oxygen and shuttled back to the matrix by the a subunit and a ring of identical c subunits across the membrane domain (FO) of ATP synthase, which is coupled to ATP synthesis. A mutation (m.9176 T > G) of the mitochondrial ATP6 gene that replaces an universally conserved leucine residue into arginine at amino acid position 217 of human subunit a (aL217R) has been associated to NARP (Neuropathy, Ataxia and Retinitis Pigmentosa) and MILS (Maternally Inherited Leigh's Syndrome) diseases. We previously showed that an equivalent thereof in Saccharomyces cerevisiae (aL237R) severely impairs subunit a assembly/stability and decreases by >90% the rate of mitochondrial ATP synthesis. Herein we identified three spontaneous first-site intragenic suppressors (aR237M, aR237T and aR237S) that fully restore ATP synthase assembly. However, mitochondrial ATP synthesis rate was only partially recovered (40-50% vs wild type yeast). In light of recently described high-resolution yeast ATP synthase structures, the detrimental consequences of the aL237R change can be explained by steric and electrostatic hindrance with the universally conserved subunit a arginine residue (aR176) that is essential to FO activity. aL237 together with three other nearby hydrophobic residues have been proposed to prevent ion shortage between two physically separated hydrophilic pockets within the FO. Our results suggest that aL237 favors subunit c-ring rotation by optimizing electrostatic interaction between aR176 and an acidic residue in subunit c (cE59) known to be essential also to the activity of FO.

Biochemical and Biophysical Research Communications, 2019

The mitochondrial ATP synthase of Polytomella exhibits a peripheral stalk and a dimerization doma... more The mitochondrial ATP synthase of Polytomella exhibits a peripheral stalk and a dimerization domain built by the Asa subunits, unique to chlorophycean algae. The topology of these subunits has been extensively studied. Here we explored the interactions of subunit Asa3 using Far Western blotting and subcomplex reconstitution, and found it associates with Asa1 and Asa8. We also identified the novel interactions Asa1-Asa2 and Asa1-Asa7. In silico analyses of Asa3 revealed that it adopts a HEAT repeat-like structure that points to its location within the enzyme based on the available 3D-map of the algal ATP synthase. We suggest that subunit Asa3 is instrumental in securing the attachment of the peripheral stalk to the membrane sector, thus stabilizing the dimeric mitochondrial ATP synthase.

Frontiers in Physiology, 2018

Rotary ATPases are a family of enzymes that are thought of as molecular nanomotors and are classi... more Rotary ATPases are a family of enzymes that are thought of as molecular nanomotors and are classified in three types: F, A, and V-type ATPases. Two members (F and A-type) can synthesize and hydrolyze ATP, depending on the energetic needs of the cell, while the V-type enzyme exhibits only a hydrolytic activity. The overall architecture of all these enzymes is conserved and three main sectors are distinguished: a catalytic core, a rotor and a stator or peripheral stalk. The peripheral stalks of the A and V-types are highly conserved in both structure and function, however, the F-type peripheral stalks have divergent structures. Furthermore, the peripheral stalk has other roles beyond its stator function, as evidenced by several biochemical and recent structural studies. This review describes the information regarding the organization of the peripheral stalk components of F, A, and V-ATPases, highlighting the key differences between the studied enzymes, as well as the different processes in which the structure is involved.

Front Physiol. 2018 Apr 4;9:329. doi: eCollection 2018., 2018

Devastating human neuromuscular disorders have been associated to defects in the ATP synthase. Th... more Devastating human neuromuscular disorders have been associated to defects in the ATP synthase. This enzyme is found in the inner mitochondrial membrane and catalyzes the last step in oxidative phosphorylation, which provides aerobic eukaryotes with ATP. With the advent of structures of complete ATP synthases, and the availability of genetically approachable systems such as the yeast Saccharomyces cerevisiae, we can begin to understand these molecular machines and their associated defects at the molecular level. In this review, we describe what is known about the clinical syndromes induced by 58 different mutations found in the mitochondrial genes encoding membrane subunits 8 and a of ATP synthase, and evaluate their functional consequences with respect to recently described cryo-EM structures.

Scientific Reports, 2016

Here we elucidated the pathogenesis of a 14-year-old Chinese female who initially developed an is... more Here we elucidated the pathogenesis of a 14-year-old Chinese female who initially developed an isolated nephropathy followed by a complex clinical presentation with brain and muscle problems, which indicated that the disease process was possibly due to a mitochondrial dysfunction. Careful evaluation of renal biopsy samples revealed a decreased staining of cells induced by COX and NADH dehydrogenase activities, and a strong fragmentation of the mitochondrial network. These anomalies were due to the presence of a mutation in the mitochondrial ATP6 gene, G8969>A. This mutation leads to replacement of a highly conserved serine residue at position 148 of the a-subunit of ATP synthase. Increasing the mutation load in cybrid cell lines was paralleled by the appearance of abnormal mitochondrial morphologies, diminished respiration and enhanced production of reactive oxygen species. An equivalent of the G8969>A mutation in yeast had dramatic consequences on ATP synthase, with a block in proton translocation. The mutation was particularly abundant (89%) in the kidney compared to blood and urine, which is likely the reason why this organ was affected first. Based on these findings, we suggest that nephrologists should pay more attention to the possibility of a mitochondrial dysfunction when evaluating patients suffering from kidney problems.

Am. J. Hum. Genet., 2016

Sudden unexpected death in infancy occurs in apparently healthy infants and remains largely unexp... more Sudden unexpected death in infancy occurs in apparently healthy infants and remains largely unexplained despite thorough investigation. The vast majority of cases are sporadic. Here we report seven individuals from three families affected by sudden and unexpected cardiac arrest between 4 and 20 months of age. Whole-exome sequencing revealed compound heterozygous missense mutations in PPA2 in affected infants of each family. PPA2 encodes the mitochondrial pyrophosphatase, which hydrolyzes inorganic pyrophosphate into two phosphates. This is an essential activity for many biosynthetic reactions and for energy metabolism of the cell. We show that deletion of the orthologous gene in yeast (ppa2Δ) compromises cell viability due to the loss of mitochondria. Expression of wild-type human PPA2, but not PPA2 containing the mutations identified in affected individuals, preserves mitochondrial function in ppa2Δ yeast. Using a regulatable (doxycycline-repressible) gene expression system, we found that the pathogenic PPA2 mutations rapidly inactivate the mitochondrial energy transducing system and prevent the maintenance of a sufficient electrical potential across the inner membrane, which explains the subsequent disappearance of mitochondria from the mutant yeast cells. Altogether these data demonstrate that PPA2 is an essential gene in yeast and that biallelic mutations in PPA2 cause a mitochondrial disease leading to sudden cardiac arrest in infants.

Since the discovery of somatic mtDNA mutations in tumor cells, multiple studies have focused on e... more Since the discovery of somatic mtDNA mutations in tumor cells, multiple studies have focused on establishing a causal relationship between those changes and alterations in energy metabolism, a hallmark of cancer cells. Yet the consequences of these mutations on mitochondrial function remain largely unknown. In this study, Saccha-romyces cerevisiae has been used as a model to investigate the functional consequences of four cancer-associated missense mutations (8914CN A, 8932CNT, 8953ANG, 9131TN C) found in the mitochondrial MT-ATP6 gene. This gene encodes the a-subunit of F 1 F O-ATP synthase, which catalyzes the last steps of ATP production in mitochon-dria. Although the four studied mutations affected well-conserved residues of the a-subunit, only one of them (8932C N T) had a significant impact on mitochondrial function, due to a less efficient incorporation of the a-subunit into ATP synthase. Our findings indicate that these ATP6 genetic variants found in human tumors are neutral mitochondrial genome substitutions with a limited, if any, impact on the energetic function of mitochondria.

The mitochondrial ATPase inhibitor, IF1, regulates the activity of the mitochondrial ATP synthase... more The mitochondrial ATPase inhibitor, IF1, regulates the activity of the mitochondrial ATP synthase. The oligomeric state of IF1 related to pH is crucial for its inhibitory activity. Although extensive structural studies have been performed to characterize the oligomeric states of bovine IF1, only little is known concerning those of yeast IF1. While bovine IF1 can be found as an inhibitory dimer at low pH and a non-inhibitory tetramer at high pH, a monomer/dimer equilibrium has been described for yeast IF1, high pH values favoring the monomeric state. Combining different strategies involving the grafting of nitroxide spin labels combined with Electron Paramag-netic Resonance (EPR) spectroscopy, the present study brings the first structural characterization, at the residue level, of yeast IF1 in its dimeric form. The results show that the dimerization interface involves the central region of the peptide revealing that the dimer corresponds to a non-inhibitory state. Moreover, we demonstrate that the C-terminal region of the peptide is highly dynamic and that this segment is probably folded back onto the central region. Finally, the pH-dependence of the inter-label distance distribution has been observed indicating a confor-mational change between two structural states in the dimer.

Mitochondrial diseases are severe and largely untreatable. Owing to the many essential processes ... more Mitochondrial diseases are severe and largely untreatable. Owing to the many essential processes carried out by mitochondria and the complex cellular systems that support these processes, these diseases are diverse, pleiotropic, and challenging to study. Much of our current understanding of mitochondrial function and dysfunction comes from studies in the baker's yeast Saccharomyces cerevisiae. Because of its good fermenting capacity, S. cerevisiae can survive mutations that inactivate oxidative phosphorylation, has the ability to tolerate the complete loss of mitochondrial DNA (a property referred to as 'petite-positivity'), and is amenable to mitochondrial and nuclear genome manipulation. These attributes make it an excellent model system for studying and resolving the molecular basis of numerous mitochondrial diseases. Here, we review the invaluable insights this model organism has yielded about diseases caused by mitochondrial dysfunction, which ranges from primary defects in oxidative phosphorylation to metabolic disorders, as well as dysfunctions in maintaining the genome or in the dynamics of mitochondria. Owing to the high level of functional conservation between yeast and human mitochondrial genes, several yeast species have been instrumental in revealing the molecular mechanisms of pathogenic human mitochondrial gene mutations. Importantly, such insights have pointed to potential therapeutic targets, as have genetic and chemical screens using yeast.

Mitochondrial F(1)F(o) ATP synthase is an enzymatic complex involved in the aerobic synthesis of ... more Mitochondrial F(1)F(o) ATP synthase is an enzymatic complex involved in the aerobic synthesis of ATP. It is well known that several enzymes are organized in supramolecular complexes in the inner mitochondrial membrane. The ATP synthase supramolecular assembly is mediated through two interfaces. One leads to dimer formation and the other to oligomer formation. In yeast, the presence of ATP synthase oligomers has been described as essential to the maintenance of the mitochondrial cristae ultrastructure. Indeed, the destabilization of the interactions between monomers was shown to alter the organization of the inner mitochondrial membrane, leading to the formation of onion-like structures similar to those observed in some mitochondrial pathologies. By using information obtained this decade (structure modeling, electron microscopy and cross-linking), this paper (i) reviews the actual state of the art and (ii) proposes a topological model of the transmembrane domains and interfaces of the ATP synthase's tetramer. This review also discusses the physiological role of this oligomerization process and its potential implications in mammal pathology. This article is part of a Directed Issue entitled: Bioenergetic Dysfunction, adaptation and therapy.

F(1)F(0) ATPases have been identified in most bacteria, including mycoplasmas which have very sma... more F(1)F(0) ATPases have been identified in most bacteria, including mycoplasmas which have very small genomes associated with a host-dependent lifestyle. In addition to the typical operon of eight genes encoding genuine F(1)F(0) ATPase (Type 1), we identified related clusters of seven genes in many mycoplasma species. Four of the encoded proteins have predicted structures similar to the α, β, γ and ε subunits of F(1) ATPases and could form an F(1)-like ATPase. The other three proteins display no similarity to any other known proteins. Two of these proteins are probably located in the membrane, as they have three and twelve predicted transmembrane helices. Phylogenomic studies identified two types of F(1)-like ATPase clusters, Type 2 and Type 3, characterized by a rapid evolution of sequences with the conservation of structural features. Clusters encoding Type 2 and Type 3 ATPases were assumed to originate from the Hominis group of mycoplasmas. We suggest that Type 3 ATPase clusters may spread to other phylogenetic groups by horizontal gene transfer between mycoplasmas in the same host, based on phylogeny and genomic context. Functional analyses in the ruminant pathogen Mycoplasma mycoides subsp. mycoides showed that the Type 3 cluster genes were organized into an operon. Proteomic analyses demonstrated that the seven encoded proteins were produced during growth in axenic media. Mutagenesis and complementation studies demonstrated an association of the Type 3 cluster with a major ATPase activity of membrane fractions. Thus, despite their tendency toward genome reduction, mycoplasmas have evolved and exchanged specific F(1)-like ATPases with no known equivalent in other bacteria. We propose a model, in which the F(1)-like structure is associated with a hypothetical X(0) sector located in the membrane of mycoplasma cells.

Journal of Biological …, Jan 1, 2011

The involvement of subunit 6 (a) in the interface between yeast ATP synthase monomers has been hi... more The involvement of subunit 6 (a) in the interface between yeast ATP synthase monomers has been highlighted. Based on the formation of a disulfide bond and using the unique cysteine 23 as target, we show that two subunits 6 are close in the inner mitochondrial membrane and in the solubilized supramolecular forms of the yeast ATP synthase. In a null mutant devoid of supernumerary subunits e and g that are involved in the stabilization of ATP synthase dimers, ATP synthase monomers are close enough in the inner mitochondrial membrane to make a disulfide bridge between their subunits 6, and this proximity is maintained in detergent extract containing this enzyme. The cross-linking of cysteine 23 located in the N-terminal part of the first transmembrane helix of subunit 6 suggests that this membrane-spanning segment is in contact with its counterpart belonging to the ATP synthase monomer that faces it and participates in the monomer-monomer interface.

… of bioenergetics and …, Jan 1, 2009

Loss of stability and integrity of large membrane protein complexes as well as their aggregation ... more Loss of stability and integrity of large membrane protein complexes as well as their aggregation in a non-lipidic environment are the major bottlenecks to their structural studies. We have tested C(12)H(25)-S-poly-Tris-(hydroxymethyl)acrylamidomethane (H(12)-TAC) among many other detergents for extracting the yeast F(1)F(0) ATP-synthase. H(12)-TAC was found to be a very efficient detergent for removing the enzyme from mitochondrial membranes without altering its sensitivity towards specific ATP-synthase inhibitors. This extracted enzyme was then solubilized by either dodecyl maltoside (DDM), H(12)-TAC or fluorinated surfactants such as C(2)H(5)-C(6)F(12)-C(2)H(4)-S-poly-Tris-(hydroxymethyl)acrylamidomethane (H(2)F(6)-TAC) or C(6)F(13)-C(2)H(4)-S-poly-Tris-(hydroxymethyl)acrylamidomethane (F(6)-TAC), two surfactants exhibiting a comparable polar head to H(12)-TAC but bearing a fluorinated hydrophobic tail. Preparations from enzymes purified in the presence of H(12)-TAC were found to be more adapted for AFM imaging than ATP-synthase purified with DDM. Keeping H(12)-TAC during the Ni-NTA IMAC purification step or replacing it by DDM at low concentrations did not however allow preserving enzyme activity, while fluorinated surfactants H(2)F(6)-TAC and F(6)-TAC were found to enhance enzyme stability and integrity as indicated by sensitivity towards inhibitors. ATPase specific activity was higher with F(6)-TAC than with H(2)F(6)-TAC. When enzymes were mixed with egg phosphatidylcholine, ATP-synthases purified in the presence of H(2)F(6)-TAC or F(6)-TAC were more stable upon time than the DDM purified enzyme. Furthermore, in the presence of lipids, an activation of ATP-synthases was observed that was transitory for enzymes purified with DDM, but lasted for weeks for ATP-synthases isolated in the presence of molecules with Tris polyalcoholic moieties. Relipidated enzymes prepared with fluorinated surfactants remained highly sensitive towards inhibitors, even after 6 weeks.

The international journal …, Jan 1, 2009

The mitochondrial F(1)F(0)-ATP synthase adopts supramolecular structures. The interaction domains... more The mitochondrial F(1)F(0)-ATP synthase adopts supramolecular structures. The interaction domains between monomers involve components belonging to the F(0) domains. In Saccharomyces cerevisiae, alteration of these components destabilizes the oligomeric structures, leading concomitantly to the appearance of monomeric species of ATP synthase and anomalous mitochondrial morphologies in the form of onion-like structures. The mitochondrial ultrastructure at the cristae level is thus modified. Electron microscopy on cross-sections of wild type mitochondria display many short cristae with narrowed intra-cristae space, whereas yeast mutants defected in supramolecular ATP synthases assembly present a low number of large lamellar cristae of constant thickness and traversing the whole organelle. The growth of these internal structures leads finally to mitochondria with sphere-like structures with a mean diameter of 1 microm that are easily identified by epifluorescence microscopy. As a result, ATP synthase is an actor of the mitochondrial ultrastructure in yeast. This paper reviews the ATP synthase components whose modifications lead to anomalous mitochondrial morphology and also provides a schema showing the formation of the so-called onion-like structures.

Atomic force microscopy (A.F.M.) was first described as a powerful technique for studying insulat... more Atomic force microscopy (A.F.M.) was first described as a powerful technique for studying insulating, hard surfaces. Since then, it has also been considered as an appropriate technique for investigating, at a submicromic scale, elastic and viscoelastic properties of soft materials as polymer films. An attempt is made to show how macroscopic models can be fruitfully employed in order to interpret the force curves obtained in AFM on polymer films. Through an analysis of the slope variation and the way the instability occurs at the tip‐sample contact, it is shown that a macroscopic approach is a useful way to explain most of the features of the force curves. Furthermore the importance is underlined of the initial conditions. It is shown that for polymer samples which have a stiffness within the range of the probe one, drastic changes of the force curve shape can occur when the initial conditions vary. Finally, this approach should allow one to clarify the conditions for which the macroscopic approach fails.

Homochiral (aMe)Leu/Ala and Leu/Ala model peptides were synthesized by solution methods and fully... more Homochiral (aMe)Leu/Ala and Leu/Ala model peptides were synthesized by solution methods and fully characterized. A solution conformational analysis of the tripeptides was carried out using FT-IR absorption and ' H NMR. The crystal-state structure of Z-D-(aMe)Leu-(D-Ala),-OMe monohydrate was resolved by X-ray diffraction. The results obtained support the conclusion that the tendency of the non-coded (aMe)Leu residue to fold into ,B-bends and helical structures is markedly higher than that of its unmethylated protein counterpart (Leu).

The X-ray diffraction crystal structures of the (αMe)Leu derivative mCIAc-d-(αMe)Leu-OH and the t... more The X-ray diffraction crystal structures of the (αMe)Leu derivative mCIAc-d-(αMe)Leu-OH and the terminally protected tripeptide Z-d-(αMe)Leu-(l-Ala)2-OMe show the onset of the fully extended (C5) conformation for the (αMe)Leu residue in both independent molecules in the asymmetric unit of the former compound and in two out of the four independent molecules in the asymmetric unit of the latter compound. In addition, conformational analysis in CDCI3 solution (using FT-infra-red absorption and 1H nuclear magnetic resonance) revealed the occurrence of a significant population of fully extended conformers throughout the entire sequence of the (αMe)Leu homochiral homopeptides pBrBz-[d-(αMe)Leu]n-OtBu (from monomer to tetramer). Taken together, these results represent a clear indication that this peptide secondary structure, uncommon for protein amino acids and other Cα-methylated chiral residues, is not a rare observation in (αMe)Leu derivatives and short peptides.

We have synthesized by solution methods and fully characterized a variety of (alpha Me)Leu/Aib mo... more We have synthesized by solution methods and fully characterized a variety of (alpha Me)Leu/Aib model peptides to the octapeptide level. A solution conformational analysis was performed by using infrared absorption. 1H nuclear magnetic resonance, and circular dichroism. The crystal-state structures of Z-D-(alpha Me)Leu-(Aib)2-OtBu, pBrBz-(Aib)2-D-(alpha Me)Leu-(Aib)2-OtBu, and Ac-(Aib)2-D-(alpha Me)Leu-(Aib)2-OtBu monohydrate were solved by x-ray diffraction. The results indicate that the (alpha Me)Leu residue may be easily incorporated into beta-bends and 3(10)-helical structures, and suggest that this residue tends to induce a helix handedness opposite to that promoted by its unmethylated counterpart (Leu) of the same optical configuration.

The crystal structure of prolyl-glutaminyl-valyl-statylalanyl-leucine (Pro-Gln-Val-Sta-Ala-Leu, C... more The crystal structure of prolyl-glutaminyl-valyl-statylalanyl-leucine (Pro-Gln-Val-Sta-Ala-Leu, C32H57N709.-5H20, Mr = 683.9 + 90.1), a putative HTLV-1 protease inhibitor based on one of the consensus retroviral protease cleavage sequences, and containing the statine residue [(4S,3S)-4-amino-3-hydroxy-6-methylheptanoic acid], has been determined by X-ray diffraction. The same molecule has been modelled in the active site of the HTLV-1 protease and both conformations have been compared. The peptide crystallizes as a pentahydrate in space group P21 with a = 10.874(2), b = 9.501(2), c = 21.062(5) A,/3 ---103.68 (1) °, Z = 2, V= 2114.3 A 3, D = 1.21 gcm -3, ~t = 8.02 cm -l, T= 293 K, A(Cu Ka) = 1.5418 A. The structure has been refined to an R value of 0.070 for 2152 observed reflections. The peptide main chain can be described as extended and adopts the usual zigzag conformation from the prolyl to the statyl residue. The main difference in conformation between the individual observed and modelled molecules is located on the Sta, Ala and Leu residues with the main chain of the modelled molecule rotated by about 180 ° as compared to the observed conformation in the crystal state.

Bovine leukaemia virus (BLV) is the aetiological agent of Leukosis enzootica bovis [Viral Oncolog... more Bovine leukaemia virus (BLV) is the aetiological agent of Leukosis enzootica bovis [Viral Oncology (1980), G. Klein (Ed.) Raven Press, New York, pp. 231-238], a widely spread disease in cattle. BLV is reported as the animal model of human T-cell leukaemia virus (HLTV) which is the causative agent of adult T-cell leukaemia and tropical spastic paraparesis. Like the viruses themselves, the two retroviral proteinases (PR) are very closely related [Virology 142 (1985) 357-377]. BLV and HTLV-I PR are reported as putative proteins made of 126 [J. Virol. 57 (1986) 826-832] and 125 [FEBS Lett. 293 (1991) 106-110] amino acids, respectively (long sequences), belonging to the aspartyl proteinase family [Nature 329 (1987) 351-354], with the aid of molecular modelling, we show that BLV and HTLV-I proteinases made of only 116 and 115 amino acids, respectively (short sequences), display three-dimensional structures similar to that observed for other retroviral aspartyl proteinases. The models are based on three-dimensional structures of Rous sarcoma virus (RSV PR) and the human immunodeficiency virus (HIV-1 PR). We used solid phase peptide synthesis to produce the putative proteolytic enzyme of BLV (116 amino acids). In this study, we show that the folded synthetic protease accurately hydrolyzes a decapeptide corresponding to the sequence of the Matrice-Capside (MA/CA) cleavage site of the gag polyprotein. In addition, the proteolytic activity is inhibited by a statine ((4S,3S)-4-amino-3-hydroxyl-6-methylheptanoic acid) containing an analogous sequence.

A complete method is presented in order to obtain from the macroscopic diffusion tensor a quantit... more A complete method is presented in order to obtain from the macroscopic diffusion tensor a quantitative microscopic model of atomic or molecular displacements. The case of self-diffusion and hetero-diffusion of naphthol-2 in single crystals of naphthalene was chosen. A vacancy mechanism with three possible molecular jumps is found to explain the observed tensor, and its anisotropy. The real jump frequencies (in the range 20 to 80 s -1 for 338 K < T < 348 K) are determined. Experimental jump energies are compared with the theoretical ones.

Molecular mobility in organic molecular crystals was studied by means of tracer diffusion experim... more Molecular mobility in organic molecular crystals was studied by means of tracer diffusion experiments: heterodiffusion at infinite dilution of 2-naphthol into naphthalene single crystals was measured at three temperatures (338, 343 and 348 K). Activation enthalpies depending on direction were obtained. The correct way to compare experimental activation energies with calculated vacancy formation and migration energies is given. A good agreement between experimental and calculated values of the activation energy of diffusion in the only direction (perpendicular to the cleavage plane) where the comparison is possible, is to be noted.

Molecular mobility in organic molecular crystal is studied by means of tracer diffusion experimen... more Molecular mobility in organic molecular crystal is studied by means of tracer diffusion experiments: hetero-diffusion at infinite dilution of 2-naphthol into naphthalene single crystals is measured at 343 K. The influence of purity on bulk diffusion is stated. The diffusion tensor is determined from measurements in different crystallographic directions. The principal axes and the anisotropy of this phenomenon are deduced.

Molecular Crystals and Liquid Crystals, Jan 1, 1986

Lattice diffusion at 343 K of β-naphthol-8 14C into naphthalene single crystals has been measured... more Lattice diffusion at 343 K of β-naphthol-8 14C into naphthalene single crystals has been measured by means of the microtome sectioning technique. Four crystallographic directions of diffusion have been studied, with two to four experiments for each of them to attain exact precision. From these data the diffusion tensor has been determined. It shows a pronounced anisotropy of the mobility of the tracer molecule into the naphthalene lattice.

A geometrical analysis of the different types of jumps into the crystalline structure of naphthalene furnishes first elements for the understanding of this anisotropy.

By means of a microtome sectioning technique, lattice diffusion at 343 K of β-naphthol-8 14C into... more By means of a microtome sectioning technique, lattice diffusion at 343 K of β-naphthol-8 14C into naphthalene single crystals has been measured in two directions. The results reveal the existence of a molecular diffusion anisotropy : Da = (12.3 ± 0.4).10-17 m2 s-1 Dc' = (4.3 ± 0.5).10-17 m2 s-1.

By means of a microtome sectioning technique, heterodiffusion coefficient of β-naphthol into naph... more By means of a microtome sectioning technique, heterodiffusion coefficient of β-naphthol into naphthalene has been measured in four directions. These results have permitted the complete determination of the diffusion tensor in this low symmetry structure and reveal the existence of a molecular diffusion anisotropy. At our knowledge it is the first complete determination of such a tensor. This anisotropy is due to a high mobility in the molecular layer than between near neighbour molecular layers

ATP (Action Thématique Programmée) Archéologie Métropolitaine, 1988

Cet article constitue la publication d’un lot de mobilier découvert en 1993 dans la commune de Ba... more Cet article constitue la publication d’un lot de mobilier découvert en 1993 dans la commune de Barbaste (Lot-et-Garonne), essentiellement composé de céramique et d’objets de parure métallique. L’étude a permis d’identifier une sépulture à incinération, probablement féminine, datable de la phase finale du premier âge du Fer aquitain, vers la fin du vie s. ou le début du ve s. a.C.

Bulletin de la Société préhistorique française, Jan 1, 1995

Documents d'archéologie …, Jan 1, 1995

Du VIe au Ier siècle av. J.-C., la vallée de la Garonne voit l'arrivée de différents produits gre... more Du VIe au Ier siècle av. J.-C., la vallée de la Garonne voit l'arrivée de différents produits grecs ou de tradition grecque. Mais le nombre d'objets ou celui des sites qui les reçoivent varient selon les périodes : expansion du VIe au IVe, déclin de la fin du IVe au IIIe siècle, reprise au IIe et au premier siècle. Du Languedoc occidental à l'estuaire girondin, la distribution du mobilier grec suit plusieurs circuits. Mais cette diffusion est-elle le résultat d'un commerce de l'étain de l'Ouest comme on l'a longtemps dit ou plutôt la preuve d'échanges diversifiés et/ou d'un colportage entre Marseille, la région languedocienne et le Sud-Ouest de la Gaule

Bulletin de la Société Préhistorique Française, Jan 1, 1983

Bulletin de la Société préhistorique …, Jan 1, 1980

Le travail présenté dans ce manuscrit a été réalisé à l'Institut de Biochimie et Génétique Cellul... more Le travail présenté dans ce manuscrit a été réalisé à l'Institut de Biochimie et Génétique Cellulaires, UMR 5095 CNRS-Université Bordeaux2, sous la direction du Professeur Ioan Lascu.

J~xprime loufe ma reconnai:5:5ance à rt/on:5ieur fe Pro/e:5:5eur Pierre f?ou9é pour m ~voir accue... more J~xprime loufe ma reconnai:5:5ance à rt/on:5ieur fe Pro/e:5:5eur Pierre f?ou9é pour m ~voir accueiiAe dan:5 :5on étjuipe . Je vou:5 remercie pour m ~voir /ail découvrir fe:5 joie:5 el fe:5 dé:5illu:5ion:5 de la recherche.

Le travail présenté dans cette thèse a été réalisé à l'Institut de Biochimie et de Génétique Cell... more Le travail présenté dans cette thèse a été réalisé à l'Institut de Biochimie et de Génétique Cellulaires du CNRS, sous la direction de Jean Velours. Je tiens à lui exprimer toute ma reconnaissance pour la formation qu'il m'a permis d'acquérir, la confiance qu'il m'a accordée et le soutien dont il m'a fait preuve dans les moments difficiles. Je remercie Monsieur le professeur Bernard Guérin pour m'avoir fait l'honneur de présider le jury de thèse. Je remercie Madame Françoise Foury, Monsieur Joël Lunardi et Monsieur Marc Le Maire pour avoir accepté d'examiner et de critiquer ce manuscrit. Je souhaite également remercier Messieurs Michel Castroviejo, Jean-Claude Gandar, Guy Lauquin et François Pénin pour leur contribution à ce travail. Merci à Christian Napias et à Stéphen Manon pour leurs précieux conseils et leur lecture attentive de ce manuscrit. Merci Geneviève pour ta gentillesse, ta disponibilité et pour m'avoir donné le goût d'enseigner. Je tiens à remercier très chaleureusement Christelle, Emmanuel, Jacques, Judith, Nathalie, Pierre-Vincent et Stéphane pour l'amitié qu'ils m'ont témoignée, pour les longues discussions scientifiques (ou non-scientifiques) que nous avons eues ensemble, pour les coups de mains qu'ils m'ont apportés et pour la bonne ambiance qu'ils ont fait régner au sein du laboratoire. Merci également à toutes les personnes des quatrième, cinquième et sixième étages pour leur sympathie et leur aide: Anne,

Présentation du travail proposé dans ce manuscrit Comment les systèmes biologiques parviennent-il... more Présentation du travail proposé dans ce manuscrit Comment les systèmes biologiques parviennent-ils à synthétiser l'ATP?

Life

International Journal of Molecular Sciences

Probing the pathogenicity and functional consequences of mitochondrial DNA (mtDNA) mutations from... more Probing the pathogenicity and functional consequences of mitochondrial DNA (mtDNA) mutations from patient’s cells and tissues is difficult due to genetic heteroplasmy (co-existence of wild type and mutated mtDNA in cells), occurrence of numerous mtDNA polymorphisms, and absence of methods for genetically transforming human mitochondria. Owing to its good fermenting capacity that enables survival to loss-of-function mtDNA mutations, its amenability to mitochondrial genome manipulation, and lack of heteroplasmy, Saccharomyces cerevisiae is an excellent model for studying and resolving the molecular bases of human diseases linked to mtDNA in a controlled genetic background. Using this model, we previously showed that a pathogenic mutation in mitochondrial ATP6 gene (m.9191T>C), that converts a highly conserved leucine residue into proline in human ATP synthase subunit a (aL222P), severely compromises the assembly of yeast ATP synthase and reduces by 90% the rate of mitochondrial ATP...

International Journal of Molecular Sciences

Bcl-xL is an oncogene of which the survival functions are finely tuned by post-translational modi... more Bcl-xL is an oncogene of which the survival functions are finely tuned by post-translational modifications (PTM). Within the Bcl-2 family of proteins, Bcl-xL shows unique eligibility to deamidation, a time-related spontaneous reaction. Deamidation is still a largely overlooked PTM due to a lack of easy techniques to monitor Asn→Asp/IsoAsp conversions or Glu→Gln conversions. Being able to detect PTMs is essential to achieve a comprehensive description of all the regulatory mechanisms and functions a protein can carry out. Here, we report a gel composition improving the electrophoretic separation of deamidated forms of Bcl-xL generated either by mutagenesis or by alkaline treatment. Importantly, this new gel formulation proved efficient to provide the long-sought evidence that even doubly-deamidated Bcl-xL remains eligible for regulation by phosphorylation.

The American Journal of Human Genetics, 2016

Sudden unexpected death in infancy occurs in apparently healthy infants and remains largely unexp... more Sudden unexpected death in infancy occurs in apparently healthy infants and remains largely unexplained despite thorough investigation. The vast majority of cases are sporadic. Here we report seven individuals from three families affected by sudden and unexpected cardiac arrest between 4 and 20 months of age. Whole-exome sequencing revealed compound heterozygous missense mutations in PPA2 in affected infants of each family. PPA2 encodes the mitochondrial pyrophosphatase, which hydrolyzes inorganic pyrophosphate into two phosphates. This is an essential activity for many biosynthetic reactions and for energy metabolism of the cell. We show that deletion of the orthologous gene in yeast (ppa2D) compromises cell viability due to the loss of mitochondria. Expression of wild-type human PPA2, but not PPA2 containing the mutations identified in affected individuals, preserves mitochondrial function in ppa2D yeast. Using a regulatable (doxycycline-repressible) gene expression system, we found that the pathogenic PPA2 mutations rapidly inactivate the mitochondrial energy transducing system and prevent the maintenance of a sufficient electrical potential across the inner membrane, which explains the subsequent disappearance of mitochondria from the mutant yeast cells. Altogether these data demonstrate that PPA2 is an essential gene in yeast and that biallelic mutations in PPA2 cause a mitochondrial disease leading to sudden cardiac arrest in infants.

European Journal of Biochemistry, 2003

A conserved putative dimerization GxxxG motif located in the unique membrane-spanning segment of ... more A conserved putative dimerization GxxxG motif located in the unique membrane-spanning segment of the ATP synthase subunit e was altered in yeast both by insertion of an alanine residue and by replacement of glycine by leucine residues. These alterations led to the loss of subunit g and the loss of dimeric and oligomeric forms of the yeast ATP synthase. Furthermore, as in null mutants devoid of either subunit e or subunit g, mitochondria displayed anomalous morphologies with onion-like structures. By taking advantage of the presence of the endogenous cysteine 28 residue in the wild-type subunit e, disulfide bond formation between subunits e in intact mitochondria was found to increase the stability of an oligomeric structure of the ATP synthase in digitonin extracts. The data show the involvement of the dimerization motif of subunit e in the formation of supramolecular structures of mitochondrial ATP synthases and are in favour of the existence in the inner mitochondrial membrane of associations of ATP synthases whose masses are higher than those of ATP synthase dimers.

Int J Biochem Cell Biol, 2009

Recueil Des Travaux Chimiques Des Pays Bas, 1994

J Pept Sci, 2002

The yeast Saccharomyces cerevisiae F 1 F O-ATPase ε-subunit (61 residues) was synthesized by the ... more The yeast Saccharomyces cerevisiae F 1 F O-ATPase ε-subunit (61 residues) was synthesized by the solid-phase peptide approach under both acidic and basic strategies. Only the latter strategy allowed us to obtain a pure ε-subunit. The strong propensity of the protein to produce few soluble dimeric species depending on pH has been proved by size-exclusion chromatography, electrophoresis and mass spectrometry. A circular dichroism study showed that an aqueous solution containing 30% trifluoroethanol or 200 mM sodium dodecyl sulphate is required for helical folding. In both solvents at acidic pH, the ε-subunit is soluble and monomeric.

Mitochondrion, Jul 12, 2016

Since the discovery of somatic mtDNA mutations in tumor cells, multiple studies have focused on e... more Since the discovery of somatic mtDNA mutations in tumor cells, multiple studies have focused on establishing a causal relationship between those changes and alterations in energy metabolism, a hallmark of cancer cells. Yet the consequences of these mutations on mitochondrial function remain largely unknown. In this study, Saccharomyces cerevisiae has been used as a model to investigate the functional consequences of four cancer-associated missense mutations (8914C>A, 8932C>T, 8953A>G, 9131T>C) found in the mitochondrial MT-ATP6 gene. This gene encodes the a-subunit of F1FO-ATP synthase, which catalyzes the last steps of ATP production in mitochondria. Although the four studied mutations affected well-conserved residues of the a-subunit, only one of them (8932C>T) had a significant impact on mitochondrial function, due to a less efficient incorporation of the a-subunit into ATP synthase. Our findings indicate that these ATP6 genetic variants found in human tumors are n...

$Research paper thumbnail of Expression, purification, crystallization and preliminary X-ray diffraction analysis of human uroporphyrinogen decarboxylase$

Acta Crystallographica Section D, May 1, 1998

Biochimica et Biophysica Acta (BBA) - Bioenergetics, 2016

The mitochondrial ATPase inhibitor, IF1, regulates the activity of the mitochondrial ATP synthase... more The mitochondrial ATPase inhibitor, IF1, regulates the activity of the mitochondrial ATP synthase. The oligomeric state of IF1 related to pH is crucial for its inhibitory activity. Although extensive structural studies have been performed to characterize the oligomeric states of bovine IF1, only little is known concerning those of yeast IF1. While bovine IF1 can be found as an inhibitory dimer at low pH and a non-inhibitory tetramer at high pH, a monomer/dimer equilibrium has been described for yeast IF1, high pH values favoring the monomeric state. Combining different strategies involving the grafting of nitroxide spin labels combined with Electron Paramagnetic Resonance (EPR) spectroscopy, the present study brings the first structural characterization, at the residue level, of yeast IF1 in its dimeric form. The results show that the dimerization interface involves the central region of the peptide revealing that the dimer corresponds to a non-inhibitory state. Moreover, we demonstrate that the C-terminal region of the peptide is highly dynamic and that this segment is probably folded back onto the central region. Finally, the pH-dependence of the inter-label distance distribution has been observed indicating a conformational change between two structural states in the dimer.

$Research paper thumbnail of Crystallization and preliminary X-ray diffraction data of mouse L-chain apoferritin crystals$

Acta Crystallographica Section D-biological Crystallography, 2000

Journal de Physique I, 1993

Disease models & mechanisms, 2015

Journal de Physique Lettres, 1985

2014 La diffusion réticulaire de 8 14C 03B2-naphtol dans le naphtalène monocristallin a été mesur... more 2014 La diffusion réticulaire de 8 14C 03B2-naphtol dans le naphtalène monocristallin a été mesurée par la méthode de sectionnement comptage dans 2 directions cristallographiques différentes. Les résultats font la preuve de l'existence d'une anisotropie de diffusion moléculaire à 343 K : Da = (12,3 ± 0,4).10-17 m2 s-1 Dc' = (4,3 ± 0,5).10-17 m2 s-1. Abstract. 2014 By means of a microtome sectioning technique, lattice diffusion at 343 K of 03B2-naphthol-8 14C into naphthalene single crystals has been measured in two directions. The results reveal the existence of a molecular diffusion anisotropy : Da = (12.3 ± 0.4).10-17 m2 s-1 Dc' = (4.3 ± 0.5).10-17 m2 s-1.

Journal de Physique, 1985

2014 On introduit une famille de modèles qui interpole entre les modèles séparables et le modèle ... more 2014 On introduit une famille de modèles qui interpole entre les modèles séparables et le modèle de Sherrington-Kirkpatrick. Ceci permet une meilleure compréhension des différences entre les modèles séparables et non séparables en particulier en ce qui concerne l'extensivité du logarithme de la fonction caractéristique des couplages aléatoires, la brisure de la symétrie des répliques et la nature des paramètres d'ordre. Cette famille contient des modèles « réalistes » comportant des paramètres ajustables susceptibles de mieux rendre compte des résultats expérimentaux que le modèle S.K. Abstract. 2014 A family of models which interpolates between the separable models and the Sherrington-Kirkpatrick (S.K.) model is introduced. This allows a better understanding of the differences between separable and non-separable models, in particular as concerns the extensivity of the logarithm of the characteristic function of the random couplings, the breaking of the replica symmetry and the nature of the order parameters. This family contains true spin glass models with adjustable parameters which might account for the experimental situation better than the S.K. model.