Emma Berta G U T I E R R E Z Cirlos | UNAM Universidad Nacional Autónoma de México (original) (raw)

Papers by Emma Berta G U T I E R R E Z Cirlos

Research paper thumbnail of The composition of the Bacillus subtilis aerobic respiratory chain supercomplexes

Journal of Bioenergetics and Biomembranes, 2012

Bacillus subtilis has a bifurcated respiratory chain composed of a cytochrome branch and a quinol... more Bacillus subtilis has a bifurcated respiratory chain composed of a cytochrome branch and a quinol oxidase branch. The respiratory complexes of this bacterium have been elucidated mostly by the analysis of the genome and by the isolation of individual complexes. The supramolecular organization of this respiratory chain is not known. In this work, we have analyzed the organization of the supercomplex in membranes isolated from B. subtilis grown in aerobic conditions in a medium with 3 % succinate. We used two different native electrophoretic techniques, clear native electrophoresis (CNE) and blue native electrophoresis (BNE). Using a heme-specific stain and Coomassie blue stain with in-gel activity assays followed by mass spectrometry, we identified the proteins resolved in both the first and second dimensions of the electrophoreses to detect the supercomplexes. We found that complexes b 6 c and caa 3 form a very high molecular mass supercomplex with the membrane-bound cytochrome c 550 and with ATP synthase. Most of the ATP synthase was found as a monomer. Succinate dehydrogenase was identified within a high molecular band between F 0 F 1 and F 1 and together with nitrate reductase. The type-2 NADH dehydrogenase was detected within a low molecular mass band. Finally, the quinol oxidase aa 3 seems to migrate as an oligomer of high molecular mass.

Research paper thumbnail of Titanium dioxide nanoparticles impair lung mitochondrial function

Toxicology Letters, 2011

Titanium dioxide nanoparticles (TiO 2 NPs) are used in an increasing number of human products suc... more Titanium dioxide nanoparticles (TiO 2 NPs) are used in an increasing number of human products such as cosmetics, sunscreen, toothpaste and paints. However, there is clear evidence about effects associated to TiO 2 NPs exposure, which include lung inflammation and tumor formation and these effects are related to reactive oxygen species (ROS) formation. The ROS generation could be attributed to a mitochondrial dysfunction. Even though, it has been shown that TiO 2 NPs exposure can induce some alterations in mitochondria including cytochrome c release to cytosol, change in mitochondrial permeability and decrease of mitochondrial membrane potential (« m), there is no information about the changes in mitochondrial function induced by TiO 2 NPs. We hypothesized that TiO 2 NPs effects are associated with mitochondrial dysfunction and redox unbalance. To test our hypothesis we isolated mitochondria from lung tissue of rats and exposed them to 10 (g TiO 2 NPs (particle size < 25 nm)/mg protein for 1 h. Our results showed that TiO 2 NPs decreases NADH levels and impairs « m and mitochondrial function accompanied by ROS generation during mitochondrial respiration.

Research paper thumbnail of Cardiolipin deficiency causes a dissociation of the b6c:caa3 megacomplex in B. subtilis membranes

Journal of Bioenergetics and Biomembranes, 2016

The associations among respiratory complexes in energy-transducing membranes have been establishe... more The associations among respiratory complexes in
energy-transducing membranes have been established. In fact,
it is known that the Gram-negative bacteria Paracoccus
denitrificans and Escherichia coli have respiratory
supercomplexes in their membranes. These supercomplexes
are important for channeling substrates between enzymes in a
metabolic pathway, and the assembly of these supercomplexes
depends on the protein subunits and membrane lipids, mainly
cardiolipin, which is present in both the mitochondrial inner
membrane and bacterial membranes. The Gram-positive bacterium
Bacillus subtilis has a branched respiratory chain, in
which some complexes generate proton motive force whereas
others constitute an escape valve of excess reducing power.
Some peculiarities of this respiratory chain are the following:
a type II NADH dehydrogenase, a unique b6c complex that
has a b6 type cytochrome with a covalently bound heme, and a
c-type heme attached to the third subunit, which is similar to
subunit IV of the photosynthetic b6f complex. Cytochrome c
oxygen reductase (caa3) contains a c-type cytochrome on subunit
I. We previously showed that the b6c and the caa3 complexes
form a supercomplex. Both the b6c and the caa3 together
with the quinol oxygen reductase aa3 generate the proton
motive force in B. subtilis. In order to seek proof that this
supercomplex is important for bacterial growth in aerobic
conditions we compared the b6c: caa3 supercomplex from
wild type membranes with membranes from two mutants lacking cardiolipin. Both mutant complexes were found to
have similar activity and heme content as the wild type. Clear
native electrophoresis showed that mutants lacking cardiolipin
had b6c:caa3 supercomplexes of lower mass or even individual
complexes after membrane solubilization with digitonin. The
use of dodecyl maltoside revealed a more evident difference
between wild-type and mutant supercomplexes. Here we
provide evidence showing that cardiolipin plays a role in the
stability of the b6c:caa3 supercomplex in B. subtilis.

Research paper thumbnail of Cardiolipin deficiency causes a dissociation of the b 6 c:caa 3 megacomplex in B. subtilis membranes

Journal of Bioenergetics and Biomembranes, 2016

The associations among respiratory complexes in energy-transducing membranes have been establishe... more The associations among respiratory complexes in energy-transducing membranes have been established. In fact, it is known that the Gram-negative bacteria Paracoccus denitrificans and Escherichia coli have respiratory supercomplexes in their membranes. These supercomplexes are important for channeling substrates between enzymes in a metabolic pathway, and the assembly of these supercomplexes depends on the protein subunits and membrane lipids, mainly cardiolipin, which is present in both the mitochondrial inner membrane and bacterial membranes. The Gram-positive bacterium Bacillus subtilis has a branched respiratory chain, in which some complexes generate proton motive force whereas others constitute an escape valve of excess reducing power. Some peculiarities of this respiratory chain are the following: a type II NADH dehydrogenase, a unique b 6 c complex that has a b 6 type cytochrome with a covalently bound heme, and a c-type heme attached to the third subunit, which is similar to subunit IV of the photosynthetic b 6 f complex. Cytochrome c oxygen reductase (caa 3 ) contains a c-type cytochrome on subunit I. We previously showed that the b 6 c and the caa 3 complexes form a supercomplex. Both the b 6 c and the caa 3 together with the quinol oxygen reductase aa 3 generate the proton motive force in B. subtilis. In order to seek proof that this supercomplex is important for bacterial growth in aerobic conditions we compared the b 6 c: caa 3 supercomplex from wild type membranes with membranes from two mutants lacking cardiolipin. Both mutant complexes were found to have similar activity and heme content as the wild type. Clear native electrophoresis showed that mutants lacking cardiolipin had b 6 c:caa 3 supercomplexes of lower mass or even individual complexes after membrane solubilization with digitonin. The use of dodecyl maltoside revealed a more evident difference between wild-type and mutant supercomplexes. Here we provide evidence showing that cardiolipin plays a role in the stability of the b 6 c:caa 3 supercomplex in B. subtilis.

Research paper thumbnail of Decrease in Respiratory Function and Electron Transport Chain Induced by Airborne Particulate Matter (PM10) Exposure in Lung Mitochondria

Toxicologic Pathology, 2013

Particulate matter, with a mean aerodynamic diameter of ≤10 µm (PM10), exposure is considered as ... more Particulate matter, with a mean aerodynamic diameter of ≤10 µm (PM10), exposure is considered as a risk factor for cardiovascular and respiratory diseases. The mechanism of cell damage induced by PM10 exposure is related to mitochondrial alterations. The aim of this work was to investigate the detailed alterations induced by PM10 on mitochondrial function. Since lung tissue is one of the most important targets of PM10 inhalation, isolated mitochondria from lung rat tissue were exposed to PM10 and structural alterations were analyzed by transmission electron microscopy. Mitochondrial function was evaluated by respiratory control index (RCI), membrane potential, adenosine triphosphate (ATP) synthesis, and activity of respiratory chain. Results showed that exposure to PM10 in isolated mitochondria from lung tissue caused enlarged intermembrane spaces and shape alterations, disruption of cristae, and the decrease in dense granules. Oxygraphic traces showed a concentration-dependent decrease in oxygen consumption and RCI. In addition, mitochondrial membrane potential, ATP synthesis, and activity of complexes II and IV showed an increase and decrease, respectively, after PM10 exposure. PM10 exposure induced disruption in structure and function in isolated mitochondria from lung rat tissue.

Research paper thumbnail of The presence of multidomain linkers determines the bundle-shape structure of the phycobilisome of the cyanobacterium Gloeobacter violaceus PCC 7421

Photosynthesis Research, 2007

The complete genome sequence of Gloeobacter violaceus [Nakamura et al. (2003a, b) DNA Res 10: [37... more The complete genome sequence of Gloeobacter violaceus [Nakamura et al. (2003a, b) DNA Res 10: [37][38][39][40][41][42][43][44][45] allows us to understand better the structure of the phycobilisomes (PBS) of this cyanobacterium. Genomic analysis revealed peculiarities in these PBS: the presence of genes for two multidomain linker proteins, a core membrane linker with four repetitive sequences (REP domains), the absence of rod core linkers, two sets of phycocyanin (PC) a and b subunits, two copies of a rod PC associated linker (CpcC), and two rod cap associated linkers (CpcD). Also, there is one ferredoxin-NADP + oxidoreductase with only two domains. The PBS proteins were investigated by gel electrophoresis, amino acid sequencing and peptide mass fingerprinting (PMF). The two unique multidomain linkers contain three REP domains with high similarity and these were found to be in tandem and were separated by dissimilar Arms. One of these, with a mass of 81 kDa, is found in heavy PBS fragments rich in PC. We propose that it links six PC hexamers in two parallel rows in the rods. The other unique linker has a mass of 91 kDa and is easily released from the heavy fragments of PBS. We propose that this links the rods to the core. The presence of these multidomain linkers could explain the bundle shaped rods of the PBS. The presence of 4 REP domains in the core membrane linker protein (129 kDa) was established by PMF. This core linker may hold together 16 AP trimers of the pentacylindrical core, or alternatively, a tetracylindrical core of the PBS of G. violaceus.

Research paper thumbnail of A proteomic approach to the analysis of the components of the phycobilisomes from two cyanobacteria with complementary chromatic adaptation: Fremyella diplosiphon UTEX B590 and Tolypothrix PCC 7601

Photosynthesis Research, 2012

Tolypothrix PCC 7601 and Fremyella diplosiphon UTEX B590 can produce two alternative phycobili-

Research paper thumbnail of Interactions of linker proteins with the phycobiliproteins in the phycobilisome substructures of Gloeobacter violaceus

Photosynthesis Research, 2010

Gloeobacter violaceus PCC 7421 is a unicellular oxygenic photosynthetic organism, which precedes ... more Gloeobacter violaceus PCC 7421 is a unicellular oxygenic photosynthetic organism, which precedes the diversification of cyanobacteria in the phylogenetic tree. It is the only cyanobacterium that does not contain internal membranes. The unique structure of the rods of the phycobilisome (PBS), grouped as one bundle of six parallel rods, distinguishes G. violaceus from the other PBS-containing cyanobacteria. It has been proposed that unique multidomain rod-linkers are responsible for this peculiarly organized shape. However, the localization of the multidomain linkers Glr1262 and Glr2806 in the PBS-rods remains controversial (Koyama et al. 2006, FEBS Lett 580:3457-3461; Krogmann et al. 2007, Photosynth Res 93:27-43). To further increase our understanding of the structure of the G. violaceus PBS, the identification of the proteins present in fractions obtained from sucrose gradient centrifugation and from native electrophoresis of partially dissociated PBS was conducted. The identification of the proteins, after electrophoresis, was done by spectrophotometry and mass spectrometry. The results support the localization of the multidomain linkers as previously proposed by us. The Glr1262 (92 kDa) linker protein was found to be the rod-core linker L(RC) (92), and Glr2806 (81 kDa), a special rod linker L(R) (81) that joins six disks of hexameric PC. Consequently, we propose to designate glr1262 as gene cpcGm (encoding L(RC) (92)) and glr2806 as gene cpcJm (encoding L(R) (81)). We also propose that the cpeC (glr1263) gene encoding L(R) (31.8) forms the interface that binds PC to PE.

Research paper thumbnail of Anti-cooperative Oxidation of Ubiquinol by the Yeast Cytochrome bc1 Complex

Journal of Biological Chemistry, 2004

We have investigated the interaction between monomers of the dimeric yeast cytochrome bc 1 comple... more We have investigated the interaction between monomers of the dimeric yeast cytochrome bc 1 complex by analyzing the pre-steady and steady state activities of the isolated enzyme in the presence of antimycin under conditions that allow the first turnover of ubiquinol oxidation to be observable in cytochrome c 1 reduction. At pH 8.8, where the redox potential of the iron-sulfur protein is ϳ200 mV and in a bc 1 complex with a mutated iron-sulfur protein of equally low redox potential, the amount of cytochrome c 1 reduced by several equivalents of decyl-ubiquinol in the presence of antimycin corresponded to only half of that present in the bc 1 complex. Similar experiments in the presence of several equivalents of cytochrome c also showed only half of the bc 1 complex participating in quinol oxidation. The extent of cytochrome b reduced corresponded to two b H hemes undergoing reduction through one center P per dimer, indicating electron transfer between the two cytochrome b subunits. Antimycin stimulated the ubiquinolcytochrome c reductase activity of the bc 1 complex at low inhibitor/enzyme ratios. This stimulation could only be fitted to a model in which half of the bc 1 dimer is inactive when both center N sites are free, becoming active upon binding of one center N inhibitor molecule per dimer, and there is electron transfer between the cytochrome b subunits of the dimer. These results are consistent with an alternating half-of-the-sites mechanism of ubiquinol oxidation in the bc 1 complex dimer.

Research paper thumbnail of Failure to Insert the Iron-Sulfur Cluster into the Rieske Iron-Sulfur Protein Impairs Both Center N and Center P of the Cytochrome bc1 Complex

Journal of Biological Chemistry, 2002

Mutation of a serine that forms a hydrogen bond to the iron-sulfur cluster of the Rieske iron-sul... more Mutation of a serine that forms a hydrogen bond to the iron-sulfur cluster of the Rieske iron-sulfur protein to a cysteine results in a respiratory-deficient yeast strain due to formation of iron-sulfur protein lacking the iron-sulfur cluster. The Rieske apoprotein lacking the iron-sulfur cluster is inserted into both monomers of the dimeric cytochrome bc(1) complex and processed to mature size, but the protein lacking iron-sulfur cluster is more susceptible to proteolysis. In addition, the protein environment of center P in one half of the dimer is affected by failure to insert the iron-sulfur cluster as indicated by the fact that only one molecule of myxothiazol can be bound to the cytochrome bc(1) dimer. Although the bc(1) complex lacking the Rieske iron-sulfur cluster cannot oxidize ubiquinol through center P, rates of reduction of cytochrome b by menaquinol through center N are normal. However, less cytochrome b is reduced through center N, and only one molecule of antimycin can be bound at center N in the bc(1) dimer lacking iron-sulfur cluster. These results indicate that failure to insert the [2Fe-2S] cluster impairs assembly of the Rieske protein into the bc(1) complex and that this interferes with proper assembly of both center P and center N in one half of the dimeric enzyme.

Research paper thumbnail of Inhibitory Analogs of Ubiquinol Act Anti-cooperatively on the Yeast Cytochrome bc1 Complex. EVIDENCE FOR AN ALTERNATING, HALF-OF-THE-SITES MECHANISM OF UBIQUINOL OXIDATION

Journal of Biological Chemistry, 2002

The cytochrome bc 1 complex is a dimeric enzyme that links electron transfer from ubiquinol to cy... more The cytochrome bc 1 complex is a dimeric enzyme that links electron transfer from ubiquinol to cytochrome c by a protonmotive Q cycle mechanism in which ubiquinol is oxidized at one center in the enzyme, referred to as center P, and ubiquinone is re-reduced at a second center, referred to as center N. To understand better the mechanism of ubiquinol oxidation, we have examined the interaction of several inhibitory analogs of ubiquinol with the yeast cytochrome bc 1 complex. Stigmatellin and methoxyacrylate stilbene, two inhibitors that block ubiquinol oxidation at center P, inhibit the yeast enzyme with a stoichiometry of 0.5 per bc 1 complex, indicating that one molecule of inhibitor is sufficient to fully inhibit the dimeric enzyme. This stoichiometry was obtained when the inhibitors were titrated in cytochrome c reductase assays and in reactions of quinol with enzyme in which the inhibitors block pre-steady state reduction of cytochrome b. As an independent measure of inhibitor binding, we titrated the red shift in the optical spectrum of ferrocytochrome b with methoxyacrylate stilbene and thus confirmed the results of the inhibition of activity titrations. The titration curves also indicate that the binding is anti-cooperative, in that a second molecule of inhibitor binds with much lower affinity to a dimer in which an inhibitor molecule is already bound. Because these inhibitors bind to the ubiquinol oxidation site in the bc 1 complex, we propose that the yeast cytochrome bc 1 complex oxidizes ubiquinol by an alternating, half-of-the-sites mechanism.

Research paper thumbnail of Inhibition of the Yeast Cytochrome bc1 Complex by Ilicicolin H, a Novel Inhibitor That Acts at the Qn Site of the bc1 Complex

Journal of Biological Chemistry, 2004

Ilicicolin H is an antibiotic isolated from the &amp;amp;amp;amp;quot;imperfect&amp;amp;a... more Ilicicolin H is an antibiotic isolated from the &amp;amp;amp;amp;quot;imperfect&amp;amp;amp;amp;quot; fungus Cylindrocladium iliciola strain MFC-870. Ilicicolin inhibits mitochondrial respiration by inhibiting the cytochrome bc(1) complex. In order to identify the site of ilicicolin action within the bc(1) complex we have characterized the effects of ilicicolin on the cytochrome bc(1) complex of Saccharomyces cerevisiae. Ilicicolin inhibits ubiquinol-cytochrome c reductase activity of the yeast bc(1) complex with an IC(50) of 3-5 nM, while 200-250 nM ilicicolin was required to obtain comparable inhibition of the bovine bc(1) complex. Ilicicolin blocks oxidation-reduction of cytochrome b through center N of the bc(1) complex and promotes oxidant-induced reduction of cytochrome b but has no effect on oxidation of ubiquinol through center P. These results indicate that ilicicolin binds to the Qn site of the bc(1) complex. Ilicicolin induces a blue shift in the absorption spectrum of ferro-cytochrome b, and titration of the spectral shift indicates binding of one inhibitor molecule per Qn site. The effects of ilicicolin on electron transfer reactions in the bc(1) complex are similar to those of antimycin, another inhibitor that binds to the Qn site of the bc(1) complex. However, because the two inhibitors have different effects on the absorption spectrum of cytochrome b, they differ in their mode of binding to the Qn site.

Research paper thumbnail of The composition of the Bacillus subtilis aerobic respiratory chain supercomplexes

Journal of Bioenergetics and Biomembranes, 2012

Bacillus subtilis has a bifurcated respiratory chain composed of a cytochrome branch and a quinol... more Bacillus subtilis has a bifurcated respiratory chain composed of a cytochrome branch and a quinol oxidase branch. The respiratory complexes of this bacterium have been elucidated mostly by the analysis of the genome and by the isolation of individual complexes. The supramolecular organization of this respiratory chain is not known. In this work, we have analyzed the organization of the supercomplex in membranes isolated from B. subtilis grown in aerobic conditions in a medium with 3 % succinate. We used two different native electrophoretic techniques, clear native electrophoresis (CNE) and blue native electrophoresis (BNE). Using a heme-specific stain and Coomassie blue stain with in-gel activity assays followed by mass spectrometry, we identified the proteins resolved in both the first and second dimensions of the electrophoreses to detect the supercomplexes. We found that complexes b ( 6 ) c and caa ( 3 ) form a very high molecular mass supercomplex with the membrane-bound cytochrome c ( 550 ) and with ATP synthase. Most of the ATP synthase was found as a monomer. Succinate dehydrogenase was identified within a high molecular band between F(0)F(1) and F(1) and together with nitrate reductase. The type-2 NADH dehydrogenase was detected within a low molecular mass band. Finally, the quinol oxidase aa ( 3 ) seems to migrate as an oligomer of high molecular mass.

Research paper thumbnail of Titanium dioxide nanoparticles impair lung mitochondrial function

Toxicology letters, Jan 25, 2011

Titanium dioxide nanoparticles (TiO(2) NPs) are used in an increasing number of human products su... more Titanium dioxide nanoparticles (TiO(2) NPs) are used in an increasing number of human products such as cosmetics, sunscreen, toothpaste and paints. However, there is clear evidence about effects associated to TiO(2) NPs exposure, which include lung inflammation and tumor formation and these effects are related to reactive oxygen species (ROS) formation. The ROS generation could be attributed to a mitochondrial dysfunction. Even though, it has been shown that TiO(2) NPs exposure can induce some alterations in mitochondria including cytochrome c release to cytosol, change in mitochondrial permeability and decrease of mitochondrial membrane potential (ΔΨ(m)), there is no information about the changes in mitochondrial function induced by TiO(2) NPs. We hypothesized that TiO(2) NPs effects are associated with mitochondrial dysfunction and redox unbalance. To test our hypothesis we isolated mitochondria from lung tissue of rats and exposed them to 10(g TiO(2) NPs (particle size<25nm)/m...

Research paper thumbnail of The phycocyanin-associated rod linker proteins of the phycobilisome of Gloeobacter violaceus PCC 7421 contain unusually located rod-capping domains

Biochimica et Biophysica Acta (BBA) - Bioenergetics, 2006

Gloeobacter violaceus PCC 7421 is a unique cyanobacterium that has no thylakoids and whose genome... more Gloeobacter violaceus PCC 7421 is a unique cyanobacterium that has no thylakoids and whose genome has been sequenced [Y. Nakamura, T. Complete Genome Structure of Gloeobacter violaceus PCC 7421, a cyanobacterium that lacks thylakoids. DNA Research 10 (2003) 137-145]. Phycobilisomes of G. violaceus were isolated and analyzed by SDS-PAGE followed by N-terminal sequencing. Three rod-linker subunits (CpeC, CpeD and CpeE) were identified as predicted from the genome sequence. The cpcC1 and cpcC2 genes at order locus named (OLN) glr0950 and gll 3219 encoding phycocyanin-associated linker proteins from G. violaceus are 56 and 55 amino acids longer at the N-terminus than the open reading frame proposed in the genome. The two amino acid extensions showed a 66% identity to one another. Also, the N-terminal extensions of these sequences were similar to domains in both the rod-capping-linker protein CpcD2 and to the C-terminus domain of the phycoerythrin-associated linker protein CpeC. These domains are not only unusual in their N-terminal location, but are unusual in that they are more closely related in sequence similarity to the C-terminus domain of the phycoerythrin-associated linker, CpeC of G. violaceus, than to the C-terminus domain of phycocyanin-associated linker CpcC in other cyanobacteria. These linker proteins with unique special domains are indicators of the unusual structure of the phycobilisomes of G. violaceus.

Research paper thumbnail of An Atypical Cytochromebin the Colorless AlgaPolytomellaspp.: The High PotentialbHHeme Exhibits a Double Transition in the α-Peak of Its Absorption Spectrum

Archives of Biochemistry and Biophysics, 1998

Polytomella spp. is a colorless alga of the family Chlamydomonadaceae that lacks chloroplasts and... more Polytomella spp. is a colorless alga of the family Chlamydomonadaceae that lacks chloroplasts and cell wall. A highly active ubiquinol-cytochrome c oxidoreductase (bc 1 complex), sensitive to antimycin and myxothiazol, has been purified and characterized from this alga (Gutié rrez-Cirlos et al., 1994, J. Biol. Chem. 269, 9147-9154). Both in mitochondrial membranes and in the isolated complex, the visible spectrum of cytochrome b from Polytomella spp. exhibits an atypical ␣-band with a maximum at 567 nm. This maximum is shifted 3-4 nm to the red when compared with b-type cytochromes from other organisms. Analysis of the b hemes of the bc 1 complex by high performance liquid chromatography revealed no differences in the retention time and in the absorption spectra of the b-type hemes from Polytomella spp. and hemin, indicating that the prosthetic group in this alga is protoheme and thus ruling out the possibility that the red-shift could be due to different chemical substitutions in the porphyrin rings of the b L or b H hemes. The two b hemes were characterized by electrochemical redox titration; at pH 7.8 -8.0, the midpoint potential for b L was ؊143 mV and for b H ؉25 mV. The spectra of the two b-type hemes were recorded in the presence of different reductants, at selected electrochemical potentials, and in the presence of antimycin A, to distinguish between the contribution of b L and b H to the visible spectrum. Both hemes b L and b H of the algal cytochrome b contribute to the observed bathochromic absorption maximum in the ␣-band of the spectrum. The data also show that the low potential b L heme from Polytomella spp. is spectroscopically similar to that of other organisms, with two transitions in the ␣-peak at 558.7 and 568.4 nm. The high-potential heme b H also exhibits a spectrum with two transitions at 557.2 and 568.9 nm, which surprisingly differs from the spectra of cytochrome b H of mammals, plants, yeasts, and bacteria, which all exhibit a single transition centered around 560 nm.

Research paper thumbnail of Extraintestinal Helminth Infection Reduces the Development of Colitis-Associated Tumorigenesis

International Journal of Biological Sciences, 2014

Colitis-associated colorectal cancer (CAC) is one of the most common cancers and is closely relat... more Colitis-associated colorectal cancer (CAC) is one of the most common cancers and is closely related to chronic or deregulated inflammation. Helminthic infections can modulate inflammatory responses in some diseases, but their immunomodulatory role during cancer development remains completely unknown. We have analyzed the role of Taenia crassiceps-induced anti-inflammatory response in determining the outcome of CAC. We show that extraintestinal T. crassiceps infection in CAC mice inhibited colonic inflammatory responses and tumor formation and prevented goblet cell loss. There was also increased expression of IL-4 and alternatively activated macrophages markers in colonic tissue and negative immunomodulation of pro-inflammatory cytokine expression. In addition, T. crassiceps infection prevented the upregulation of β-catenin and CXCR2 expression observed in the CAC mice, which are both markers associated with CAC-tumorigenesis, and reduced the numbers of circulating and colonic CD11b(+)Ly6C(hi)CCR2(+) monocytes. Thus, immunomodulatory activities induced by helminth infections may have a role in the progression of CAC.

Research paper thumbnail of The composition of the Bacillus subtilis aerobic respiratory chain supercomplexes

Journal of Bioenergetics and Biomembranes, 2012

Bacillus subtilis has a bifurcated respiratory chain composed of a cytochrome branch and a quinol... more Bacillus subtilis has a bifurcated respiratory chain composed of a cytochrome branch and a quinol oxidase branch. The respiratory complexes of this bacterium have been elucidated mostly by the analysis of the genome and by the isolation of individual complexes. The supramolecular organization of this respiratory chain is not known. In this work, we have analyzed the organization of the supercomplex in membranes isolated from B. subtilis grown in aerobic conditions in a medium with 3 % succinate. We used two different native electrophoretic techniques, clear native electrophoresis (CNE) and blue native electrophoresis (BNE). Using a heme-specific stain and Coomassie blue stain with in-gel activity assays followed by mass spectrometry, we identified the proteins resolved in both the first and second dimensions of the electrophoreses to detect the supercomplexes. We found that complexes b 6 c and caa 3 form a very high molecular mass supercomplex with the membrane-bound cytochrome c 550 and with ATP synthase. Most of the ATP synthase was found as a monomer. Succinate dehydrogenase was identified within a high molecular band between F 0 F 1 and F 1 and together with nitrate reductase. The type-2 NADH dehydrogenase was detected within a low molecular mass band. Finally, the quinol oxidase aa 3 seems to migrate as an oligomer of high molecular mass.

Research paper thumbnail of Titanium dioxide nanoparticles impair lung mitochondrial function

Toxicology Letters, 2011

Titanium dioxide nanoparticles (TiO 2 NPs) are used in an increasing number of human products suc... more Titanium dioxide nanoparticles (TiO 2 NPs) are used in an increasing number of human products such as cosmetics, sunscreen, toothpaste and paints. However, there is clear evidence about effects associated to TiO 2 NPs exposure, which include lung inflammation and tumor formation and these effects are related to reactive oxygen species (ROS) formation. The ROS generation could be attributed to a mitochondrial dysfunction. Even though, it has been shown that TiO 2 NPs exposure can induce some alterations in mitochondria including cytochrome c release to cytosol, change in mitochondrial permeability and decrease of mitochondrial membrane potential (« m), there is no information about the changes in mitochondrial function induced by TiO 2 NPs. We hypothesized that TiO 2 NPs effects are associated with mitochondrial dysfunction and redox unbalance. To test our hypothesis we isolated mitochondria from lung tissue of rats and exposed them to 10 (g TiO 2 NPs (particle size < 25 nm)/mg protein for 1 h. Our results showed that TiO 2 NPs decreases NADH levels and impairs « m and mitochondrial function accompanied by ROS generation during mitochondrial respiration.

Research paper thumbnail of Cardiolipin deficiency causes a dissociation of the b6c:caa3 megacomplex in B. subtilis membranes

Journal of Bioenergetics and Biomembranes, 2016

The associations among respiratory complexes in energy-transducing membranes have been establishe... more The associations among respiratory complexes in
energy-transducing membranes have been established. In fact,
it is known that the Gram-negative bacteria Paracoccus
denitrificans and Escherichia coli have respiratory
supercomplexes in their membranes. These supercomplexes
are important for channeling substrates between enzymes in a
metabolic pathway, and the assembly of these supercomplexes
depends on the protein subunits and membrane lipids, mainly
cardiolipin, which is present in both the mitochondrial inner
membrane and bacterial membranes. The Gram-positive bacterium
Bacillus subtilis has a branched respiratory chain, in
which some complexes generate proton motive force whereas
others constitute an escape valve of excess reducing power.
Some peculiarities of this respiratory chain are the following:
a type II NADH dehydrogenase, a unique b6c complex that
has a b6 type cytochrome with a covalently bound heme, and a
c-type heme attached to the third subunit, which is similar to
subunit IV of the photosynthetic b6f complex. Cytochrome c
oxygen reductase (caa3) contains a c-type cytochrome on subunit
I. We previously showed that the b6c and the caa3 complexes
form a supercomplex. Both the b6c and the caa3 together
with the quinol oxygen reductase aa3 generate the proton
motive force in B. subtilis. In order to seek proof that this
supercomplex is important for bacterial growth in aerobic
conditions we compared the b6c: caa3 supercomplex from
wild type membranes with membranes from two mutants lacking cardiolipin. Both mutant complexes were found to
have similar activity and heme content as the wild type. Clear
native electrophoresis showed that mutants lacking cardiolipin
had b6c:caa3 supercomplexes of lower mass or even individual
complexes after membrane solubilization with digitonin. The
use of dodecyl maltoside revealed a more evident difference
between wild-type and mutant supercomplexes. Here we
provide evidence showing that cardiolipin plays a role in the
stability of the b6c:caa3 supercomplex in B. subtilis.

Research paper thumbnail of Cardiolipin deficiency causes a dissociation of the b 6 c:caa 3 megacomplex in B. subtilis membranes

Journal of Bioenergetics and Biomembranes, 2016

The associations among respiratory complexes in energy-transducing membranes have been establishe... more The associations among respiratory complexes in energy-transducing membranes have been established. In fact, it is known that the Gram-negative bacteria Paracoccus denitrificans and Escherichia coli have respiratory supercomplexes in their membranes. These supercomplexes are important for channeling substrates between enzymes in a metabolic pathway, and the assembly of these supercomplexes depends on the protein subunits and membrane lipids, mainly cardiolipin, which is present in both the mitochondrial inner membrane and bacterial membranes. The Gram-positive bacterium Bacillus subtilis has a branched respiratory chain, in which some complexes generate proton motive force whereas others constitute an escape valve of excess reducing power. Some peculiarities of this respiratory chain are the following: a type II NADH dehydrogenase, a unique b 6 c complex that has a b 6 type cytochrome with a covalently bound heme, and a c-type heme attached to the third subunit, which is similar to subunit IV of the photosynthetic b 6 f complex. Cytochrome c oxygen reductase (caa 3 ) contains a c-type cytochrome on subunit I. We previously showed that the b 6 c and the caa 3 complexes form a supercomplex. Both the b 6 c and the caa 3 together with the quinol oxygen reductase aa 3 generate the proton motive force in B. subtilis. In order to seek proof that this supercomplex is important for bacterial growth in aerobic conditions we compared the b 6 c: caa 3 supercomplex from wild type membranes with membranes from two mutants lacking cardiolipin. Both mutant complexes were found to have similar activity and heme content as the wild type. Clear native electrophoresis showed that mutants lacking cardiolipin had b 6 c:caa 3 supercomplexes of lower mass or even individual complexes after membrane solubilization with digitonin. The use of dodecyl maltoside revealed a more evident difference between wild-type and mutant supercomplexes. Here we provide evidence showing that cardiolipin plays a role in the stability of the b 6 c:caa 3 supercomplex in B. subtilis.

Research paper thumbnail of Decrease in Respiratory Function and Electron Transport Chain Induced by Airborne Particulate Matter (PM10) Exposure in Lung Mitochondria

Toxicologic Pathology, 2013

Particulate matter, with a mean aerodynamic diameter of ≤10 µm (PM10), exposure is considered as ... more Particulate matter, with a mean aerodynamic diameter of ≤10 µm (PM10), exposure is considered as a risk factor for cardiovascular and respiratory diseases. The mechanism of cell damage induced by PM10 exposure is related to mitochondrial alterations. The aim of this work was to investigate the detailed alterations induced by PM10 on mitochondrial function. Since lung tissue is one of the most important targets of PM10 inhalation, isolated mitochondria from lung rat tissue were exposed to PM10 and structural alterations were analyzed by transmission electron microscopy. Mitochondrial function was evaluated by respiratory control index (RCI), membrane potential, adenosine triphosphate (ATP) synthesis, and activity of respiratory chain. Results showed that exposure to PM10 in isolated mitochondria from lung tissue caused enlarged intermembrane spaces and shape alterations, disruption of cristae, and the decrease in dense granules. Oxygraphic traces showed a concentration-dependent decrease in oxygen consumption and RCI. In addition, mitochondrial membrane potential, ATP synthesis, and activity of complexes II and IV showed an increase and decrease, respectively, after PM10 exposure. PM10 exposure induced disruption in structure and function in isolated mitochondria from lung rat tissue.

Research paper thumbnail of The presence of multidomain linkers determines the bundle-shape structure of the phycobilisome of the cyanobacterium Gloeobacter violaceus PCC 7421

Photosynthesis Research, 2007

The complete genome sequence of Gloeobacter violaceus [Nakamura et al. (2003a, b) DNA Res 10: [37... more The complete genome sequence of Gloeobacter violaceus [Nakamura et al. (2003a, b) DNA Res 10: [37][38][39][40][41][42][43][44][45] allows us to understand better the structure of the phycobilisomes (PBS) of this cyanobacterium. Genomic analysis revealed peculiarities in these PBS: the presence of genes for two multidomain linker proteins, a core membrane linker with four repetitive sequences (REP domains), the absence of rod core linkers, two sets of phycocyanin (PC) a and b subunits, two copies of a rod PC associated linker (CpcC), and two rod cap associated linkers (CpcD). Also, there is one ferredoxin-NADP + oxidoreductase with only two domains. The PBS proteins were investigated by gel electrophoresis, amino acid sequencing and peptide mass fingerprinting (PMF). The two unique multidomain linkers contain three REP domains with high similarity and these were found to be in tandem and were separated by dissimilar Arms. One of these, with a mass of 81 kDa, is found in heavy PBS fragments rich in PC. We propose that it links six PC hexamers in two parallel rows in the rods. The other unique linker has a mass of 91 kDa and is easily released from the heavy fragments of PBS. We propose that this links the rods to the core. The presence of these multidomain linkers could explain the bundle shaped rods of the PBS. The presence of 4 REP domains in the core membrane linker protein (129 kDa) was established by PMF. This core linker may hold together 16 AP trimers of the pentacylindrical core, or alternatively, a tetracylindrical core of the PBS of G. violaceus.

Research paper thumbnail of A proteomic approach to the analysis of the components of the phycobilisomes from two cyanobacteria with complementary chromatic adaptation: Fremyella diplosiphon UTEX B590 and Tolypothrix PCC 7601

Photosynthesis Research, 2012

Tolypothrix PCC 7601 and Fremyella diplosiphon UTEX B590 can produce two alternative phycobili-

Research paper thumbnail of Interactions of linker proteins with the phycobiliproteins in the phycobilisome substructures of Gloeobacter violaceus

Photosynthesis Research, 2010

Gloeobacter violaceus PCC 7421 is a unicellular oxygenic photosynthetic organism, which precedes ... more Gloeobacter violaceus PCC 7421 is a unicellular oxygenic photosynthetic organism, which precedes the diversification of cyanobacteria in the phylogenetic tree. It is the only cyanobacterium that does not contain internal membranes. The unique structure of the rods of the phycobilisome (PBS), grouped as one bundle of six parallel rods, distinguishes G. violaceus from the other PBS-containing cyanobacteria. It has been proposed that unique multidomain rod-linkers are responsible for this peculiarly organized shape. However, the localization of the multidomain linkers Glr1262 and Glr2806 in the PBS-rods remains controversial (Koyama et al. 2006, FEBS Lett 580:3457-3461; Krogmann et al. 2007, Photosynth Res 93:27-43). To further increase our understanding of the structure of the G. violaceus PBS, the identification of the proteins present in fractions obtained from sucrose gradient centrifugation and from native electrophoresis of partially dissociated PBS was conducted. The identification of the proteins, after electrophoresis, was done by spectrophotometry and mass spectrometry. The results support the localization of the multidomain linkers as previously proposed by us. The Glr1262 (92 kDa) linker protein was found to be the rod-core linker L(RC) (92), and Glr2806 (81 kDa), a special rod linker L(R) (81) that joins six disks of hexameric PC. Consequently, we propose to designate glr1262 as gene cpcGm (encoding L(RC) (92)) and glr2806 as gene cpcJm (encoding L(R) (81)). We also propose that the cpeC (glr1263) gene encoding L(R) (31.8) forms the interface that binds PC to PE.

Research paper thumbnail of Anti-cooperative Oxidation of Ubiquinol by the Yeast Cytochrome bc1 Complex

Journal of Biological Chemistry, 2004

We have investigated the interaction between monomers of the dimeric yeast cytochrome bc 1 comple... more We have investigated the interaction between monomers of the dimeric yeast cytochrome bc 1 complex by analyzing the pre-steady and steady state activities of the isolated enzyme in the presence of antimycin under conditions that allow the first turnover of ubiquinol oxidation to be observable in cytochrome c 1 reduction. At pH 8.8, where the redox potential of the iron-sulfur protein is ϳ200 mV and in a bc 1 complex with a mutated iron-sulfur protein of equally low redox potential, the amount of cytochrome c 1 reduced by several equivalents of decyl-ubiquinol in the presence of antimycin corresponded to only half of that present in the bc 1 complex. Similar experiments in the presence of several equivalents of cytochrome c also showed only half of the bc 1 complex participating in quinol oxidation. The extent of cytochrome b reduced corresponded to two b H hemes undergoing reduction through one center P per dimer, indicating electron transfer between the two cytochrome b subunits. Antimycin stimulated the ubiquinolcytochrome c reductase activity of the bc 1 complex at low inhibitor/enzyme ratios. This stimulation could only be fitted to a model in which half of the bc 1 dimer is inactive when both center N sites are free, becoming active upon binding of one center N inhibitor molecule per dimer, and there is electron transfer between the cytochrome b subunits of the dimer. These results are consistent with an alternating half-of-the-sites mechanism of ubiquinol oxidation in the bc 1 complex dimer.

Research paper thumbnail of Failure to Insert the Iron-Sulfur Cluster into the Rieske Iron-Sulfur Protein Impairs Both Center N and Center P of the Cytochrome bc1 Complex

Journal of Biological Chemistry, 2002

Mutation of a serine that forms a hydrogen bond to the iron-sulfur cluster of the Rieske iron-sul... more Mutation of a serine that forms a hydrogen bond to the iron-sulfur cluster of the Rieske iron-sulfur protein to a cysteine results in a respiratory-deficient yeast strain due to formation of iron-sulfur protein lacking the iron-sulfur cluster. The Rieske apoprotein lacking the iron-sulfur cluster is inserted into both monomers of the dimeric cytochrome bc(1) complex and processed to mature size, but the protein lacking iron-sulfur cluster is more susceptible to proteolysis. In addition, the protein environment of center P in one half of the dimer is affected by failure to insert the iron-sulfur cluster as indicated by the fact that only one molecule of myxothiazol can be bound to the cytochrome bc(1) dimer. Although the bc(1) complex lacking the Rieske iron-sulfur cluster cannot oxidize ubiquinol through center P, rates of reduction of cytochrome b by menaquinol through center N are normal. However, less cytochrome b is reduced through center N, and only one molecule of antimycin can be bound at center N in the bc(1) dimer lacking iron-sulfur cluster. These results indicate that failure to insert the [2Fe-2S] cluster impairs assembly of the Rieske protein into the bc(1) complex and that this interferes with proper assembly of both center P and center N in one half of the dimeric enzyme.

Research paper thumbnail of Inhibitory Analogs of Ubiquinol Act Anti-cooperatively on the Yeast Cytochrome bc1 Complex. EVIDENCE FOR AN ALTERNATING, HALF-OF-THE-SITES MECHANISM OF UBIQUINOL OXIDATION

Journal of Biological Chemistry, 2002

The cytochrome bc 1 complex is a dimeric enzyme that links electron transfer from ubiquinol to cy... more The cytochrome bc 1 complex is a dimeric enzyme that links electron transfer from ubiquinol to cytochrome c by a protonmotive Q cycle mechanism in which ubiquinol is oxidized at one center in the enzyme, referred to as center P, and ubiquinone is re-reduced at a second center, referred to as center N. To understand better the mechanism of ubiquinol oxidation, we have examined the interaction of several inhibitory analogs of ubiquinol with the yeast cytochrome bc 1 complex. Stigmatellin and methoxyacrylate stilbene, two inhibitors that block ubiquinol oxidation at center P, inhibit the yeast enzyme with a stoichiometry of 0.5 per bc 1 complex, indicating that one molecule of inhibitor is sufficient to fully inhibit the dimeric enzyme. This stoichiometry was obtained when the inhibitors were titrated in cytochrome c reductase assays and in reactions of quinol with enzyme in which the inhibitors block pre-steady state reduction of cytochrome b. As an independent measure of inhibitor binding, we titrated the red shift in the optical spectrum of ferrocytochrome b with methoxyacrylate stilbene and thus confirmed the results of the inhibition of activity titrations. The titration curves also indicate that the binding is anti-cooperative, in that a second molecule of inhibitor binds with much lower affinity to a dimer in which an inhibitor molecule is already bound. Because these inhibitors bind to the ubiquinol oxidation site in the bc 1 complex, we propose that the yeast cytochrome bc 1 complex oxidizes ubiquinol by an alternating, half-of-the-sites mechanism.

Research paper thumbnail of Inhibition of the Yeast Cytochrome bc1 Complex by Ilicicolin H, a Novel Inhibitor That Acts at the Qn Site of the bc1 Complex

Journal of Biological Chemistry, 2004

Ilicicolin H is an antibiotic isolated from the &amp;amp;amp;amp;quot;imperfect&amp;amp;a... more Ilicicolin H is an antibiotic isolated from the &amp;amp;amp;amp;quot;imperfect&amp;amp;amp;amp;quot; fungus Cylindrocladium iliciola strain MFC-870. Ilicicolin inhibits mitochondrial respiration by inhibiting the cytochrome bc(1) complex. In order to identify the site of ilicicolin action within the bc(1) complex we have characterized the effects of ilicicolin on the cytochrome bc(1) complex of Saccharomyces cerevisiae. Ilicicolin inhibits ubiquinol-cytochrome c reductase activity of the yeast bc(1) complex with an IC(50) of 3-5 nM, while 200-250 nM ilicicolin was required to obtain comparable inhibition of the bovine bc(1) complex. Ilicicolin blocks oxidation-reduction of cytochrome b through center N of the bc(1) complex and promotes oxidant-induced reduction of cytochrome b but has no effect on oxidation of ubiquinol through center P. These results indicate that ilicicolin binds to the Qn site of the bc(1) complex. Ilicicolin induces a blue shift in the absorption spectrum of ferro-cytochrome b, and titration of the spectral shift indicates binding of one inhibitor molecule per Qn site. The effects of ilicicolin on electron transfer reactions in the bc(1) complex are similar to those of antimycin, another inhibitor that binds to the Qn site of the bc(1) complex. However, because the two inhibitors have different effects on the absorption spectrum of cytochrome b, they differ in their mode of binding to the Qn site.

Research paper thumbnail of The composition of the Bacillus subtilis aerobic respiratory chain supercomplexes

Journal of Bioenergetics and Biomembranes, 2012

Bacillus subtilis has a bifurcated respiratory chain composed of a cytochrome branch and a quinol... more Bacillus subtilis has a bifurcated respiratory chain composed of a cytochrome branch and a quinol oxidase branch. The respiratory complexes of this bacterium have been elucidated mostly by the analysis of the genome and by the isolation of individual complexes. The supramolecular organization of this respiratory chain is not known. In this work, we have analyzed the organization of the supercomplex in membranes isolated from B. subtilis grown in aerobic conditions in a medium with 3 % succinate. We used two different native electrophoretic techniques, clear native electrophoresis (CNE) and blue native electrophoresis (BNE). Using a heme-specific stain and Coomassie blue stain with in-gel activity assays followed by mass spectrometry, we identified the proteins resolved in both the first and second dimensions of the electrophoreses to detect the supercomplexes. We found that complexes b ( 6 ) c and caa ( 3 ) form a very high molecular mass supercomplex with the membrane-bound cytochrome c ( 550 ) and with ATP synthase. Most of the ATP synthase was found as a monomer. Succinate dehydrogenase was identified within a high molecular band between F(0)F(1) and F(1) and together with nitrate reductase. The type-2 NADH dehydrogenase was detected within a low molecular mass band. Finally, the quinol oxidase aa ( 3 ) seems to migrate as an oligomer of high molecular mass.

Research paper thumbnail of Titanium dioxide nanoparticles impair lung mitochondrial function

Toxicology letters, Jan 25, 2011

Titanium dioxide nanoparticles (TiO(2) NPs) are used in an increasing number of human products su... more Titanium dioxide nanoparticles (TiO(2) NPs) are used in an increasing number of human products such as cosmetics, sunscreen, toothpaste and paints. However, there is clear evidence about effects associated to TiO(2) NPs exposure, which include lung inflammation and tumor formation and these effects are related to reactive oxygen species (ROS) formation. The ROS generation could be attributed to a mitochondrial dysfunction. Even though, it has been shown that TiO(2) NPs exposure can induce some alterations in mitochondria including cytochrome c release to cytosol, change in mitochondrial permeability and decrease of mitochondrial membrane potential (ΔΨ(m)), there is no information about the changes in mitochondrial function induced by TiO(2) NPs. We hypothesized that TiO(2) NPs effects are associated with mitochondrial dysfunction and redox unbalance. To test our hypothesis we isolated mitochondria from lung tissue of rats and exposed them to 10(g TiO(2) NPs (particle size<25nm)/m...

Research paper thumbnail of The phycocyanin-associated rod linker proteins of the phycobilisome of Gloeobacter violaceus PCC 7421 contain unusually located rod-capping domains

Biochimica et Biophysica Acta (BBA) - Bioenergetics, 2006

Gloeobacter violaceus PCC 7421 is a unique cyanobacterium that has no thylakoids and whose genome... more Gloeobacter violaceus PCC 7421 is a unique cyanobacterium that has no thylakoids and whose genome has been sequenced [Y. Nakamura, T. Complete Genome Structure of Gloeobacter violaceus PCC 7421, a cyanobacterium that lacks thylakoids. DNA Research 10 (2003) 137-145]. Phycobilisomes of G. violaceus were isolated and analyzed by SDS-PAGE followed by N-terminal sequencing. Three rod-linker subunits (CpeC, CpeD and CpeE) were identified as predicted from the genome sequence. The cpcC1 and cpcC2 genes at order locus named (OLN) glr0950 and gll 3219 encoding phycocyanin-associated linker proteins from G. violaceus are 56 and 55 amino acids longer at the N-terminus than the open reading frame proposed in the genome. The two amino acid extensions showed a 66% identity to one another. Also, the N-terminal extensions of these sequences were similar to domains in both the rod-capping-linker protein CpcD2 and to the C-terminus domain of the phycoerythrin-associated linker protein CpeC. These domains are not only unusual in their N-terminal location, but are unusual in that they are more closely related in sequence similarity to the C-terminus domain of the phycoerythrin-associated linker, CpeC of G. violaceus, than to the C-terminus domain of phycocyanin-associated linker CpcC in other cyanobacteria. These linker proteins with unique special domains are indicators of the unusual structure of the phycobilisomes of G. violaceus.

Research paper thumbnail of An Atypical Cytochromebin the Colorless AlgaPolytomellaspp.: The High PotentialbHHeme Exhibits a Double Transition in the α-Peak of Its Absorption Spectrum

Archives of Biochemistry and Biophysics, 1998

Polytomella spp. is a colorless alga of the family Chlamydomonadaceae that lacks chloroplasts and... more Polytomella spp. is a colorless alga of the family Chlamydomonadaceae that lacks chloroplasts and cell wall. A highly active ubiquinol-cytochrome c oxidoreductase (bc 1 complex), sensitive to antimycin and myxothiazol, has been purified and characterized from this alga (Gutié rrez-Cirlos et al., 1994, J. Biol. Chem. 269, 9147-9154). Both in mitochondrial membranes and in the isolated complex, the visible spectrum of cytochrome b from Polytomella spp. exhibits an atypical ␣-band with a maximum at 567 nm. This maximum is shifted 3-4 nm to the red when compared with b-type cytochromes from other organisms. Analysis of the b hemes of the bc 1 complex by high performance liquid chromatography revealed no differences in the retention time and in the absorption spectra of the b-type hemes from Polytomella spp. and hemin, indicating that the prosthetic group in this alga is protoheme and thus ruling out the possibility that the red-shift could be due to different chemical substitutions in the porphyrin rings of the b L or b H hemes. The two b hemes were characterized by electrochemical redox titration; at pH 7.8 -8.0, the midpoint potential for b L was ؊143 mV and for b H ؉25 mV. The spectra of the two b-type hemes were recorded in the presence of different reductants, at selected electrochemical potentials, and in the presence of antimycin A, to distinguish between the contribution of b L and b H to the visible spectrum. Both hemes b L and b H of the algal cytochrome b contribute to the observed bathochromic absorption maximum in the ␣-band of the spectrum. The data also show that the low potential b L heme from Polytomella spp. is spectroscopically similar to that of other organisms, with two transitions in the ␣-peak at 558.7 and 568.4 nm. The high-potential heme b H also exhibits a spectrum with two transitions at 557.2 and 568.9 nm, which surprisingly differs from the spectra of cytochrome b H of mammals, plants, yeasts, and bacteria, which all exhibit a single transition centered around 560 nm.

Research paper thumbnail of Extraintestinal Helminth Infection Reduces the Development of Colitis-Associated Tumorigenesis

International Journal of Biological Sciences, 2014

Colitis-associated colorectal cancer (CAC) is one of the most common cancers and is closely relat... more Colitis-associated colorectal cancer (CAC) is one of the most common cancers and is closely related to chronic or deregulated inflammation. Helminthic infections can modulate inflammatory responses in some diseases, but their immunomodulatory role during cancer development remains completely unknown. We have analyzed the role of Taenia crassiceps-induced anti-inflammatory response in determining the outcome of CAC. We show that extraintestinal T. crassiceps infection in CAC mice inhibited colonic inflammatory responses and tumor formation and prevented goblet cell loss. There was also increased expression of IL-4 and alternatively activated macrophages markers in colonic tissue and negative immunomodulation of pro-inflammatory cytokine expression. In addition, T. crassiceps infection prevented the upregulation of β-catenin and CXCR2 expression observed in the CAC mice, which are both markers associated with CAC-tumorigenesis, and reduced the numbers of circulating and colonic CD11b(+)Ly6C(hi)CCR2(+) monocytes. Thus, immunomodulatory activities induced by helminth infections may have a role in the progression of CAC.