Alexander Dikiy - Academia.edu (original) (raw)

Papers by Alexander Dikiy

Biochimica Et Biophysica Acta-protein Structure and Molecular Enzymology, 2000

Cytochrome c from the methylotrophic yeast Hansenula polymorpha was isolated and purified to homo... more Cytochrome c from the methylotrophic yeast Hansenula polymorpha was isolated and purified to homogeneity for the first time. The final yield of the highly purified protein from 1.4 kg (wet weight) cells was about 20 mg. The hemoprotein has an apparent molecular mass of 12 kDa and isoelectric point (pI) of 9.3. The purified protein was characterized by electronic, EPR

Advances in Photosynthesis and Respiration, 2014

Journal of Magnetic Resonance, Series B, 1994

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PLoS ONE, 2012

Thioredoxin glutathione reductase (TGR) is a member of the mammalian thioredoxin reductase family... more Thioredoxin glutathione reductase (TGR) is a member of the mammalian thioredoxin reductase family that has a monothiol glutaredoxin (Grx) domain attached to the thioredoxin reductase module. Here, we report a structure of the Grx domain of mouse TGR, determined through high resolution NMR spectroscopy to the final backbone RMSD value of 0.4860.10 Å . The structure represents a sandwich-like molecule composed of a four stranded b-sheet flanked by five a-helixes, with the CxxS active motif located on the catalytic loop. We structurally characterized the glutathione-binding site in the protein and describe sequence and structural relationships of the domain with glutaredoxins. The structure illuminates a key functional center that evolved in mammalian TGRs to act in thiol-disulfide reactions. Our study allows us to hypothesize that Cys105 might be functionally relevant for TGR catalysis. In addition, the data suggest that the N-terminus of Grx acts as a possible regulatory signal also protecting the protein active site from unwanted interactions in cellular cytosol. Citation: Dobrovolska O, Shumilina E, Gladyshev VN, Dikiy A (2012) Structural Analysis of Glutaredoxin Domain of Mus musculus Thioredoxin Glutathione Reductase. PLoS ONE 7(12): e52914.

Selenium, 2011

... Harvard Medical School , Boston , MA 02115 , USA A. Dikiy (*) Department of Biotechnology , N... more ... Harvard Medical School , Boston , MA 02115 , USA A. Dikiy (*) Department of Biotechnology , Norwegian University of Science and Technology , 7491 Trondheim , Norway e-mail:alex.dikiy@biotech ... Dikiy A, Novoselov SV, Fomenko DE et al (2007) Biochemistry 46:6871 29. ...

Biomolecular NMR Assignments, 2007

Isotopically labeled, 15N and 15N/13C forms of recombinant methionine-r-sulfoxide reductase 1 (Ms... more Isotopically labeled, 15N and 15N/13C forms of recombinant methionine-r-sulfoxide reductase 1 (MsrB1, SelR) from Mus musculus were produced, in which catalytic selenocysteine was replaced with cysteine. We report here the 1H, 13C and 15N NMR assignment of the reduced form of this mammalian protein.

Biomolecular NMR Assignments, 2011

Two forms of the glutaredoxin (Grx) domain (full length Grx domain and short Grx lacking the Nter... more Two forms of the glutaredoxin (Grx) domain (full length Grx domain and short Grx lacking the Nterminal region) of Mus musculus thioredoxin glutathione reductase (TGR) were isotopically labelled with 15 N and 13 C isotopes, expressed and purified to homogeneity. We report here the 1 H, 13 C and 15 N NMR assignment for both Grx forms of this mouse TGR. This investigation represents the first NMR analysis of a mammalian TGR.

Biomolecular NMR Assignments, 2008

A recombinant mouse methionine-R-sulfoxide reductase 2 (MsrB2ΔS) isotopically labeled with 15 N a... more A recombinant mouse methionine-R-sulfoxide reductase 2 (MsrB2ΔS) isotopically labeled with 15 N and 15 N/ 13 C was generated. We report here the 1 H, 15 N and 13 C NMR assignments of the reduced form of this protein.

Proteins, 2011

Methionine sulfoxide reductases are antioxidant enzymes that repair oxidatively damaged methionin... more Methionine sulfoxide reductases are antioxidant enzymes that repair oxidatively damaged methionine residues in proteins. Mammals have three members of the methionine-R-sulfoxide reductase family, including cytosolic MsrB1, mitochondrial MsrB2, and endoplasmic reticulum MsrB3. Here, we report the solution structure of reduced Mus musculus MsrB2 using high resolution nuclear magnetic resonance (NMR) spectroscopy. MsrB2 is a β-strand rich globular protein consisting of eight antiparallel β-strands and three N-terminal α-helical segments. The latter secondary structure elements represent the main structural difference between mammalian MsrB2 and MsrB1. Structural comparison of mammalian and bacterial MsrB structures indicates that the general topology of this MsrB family is maintained and that MsrB2 more resembles bacterial MsrBs than MsrB1. Structural and biochemical analysis supports the catalytic mechanism of MsrB2 that, in contrast to MsrB1, does not involve a resolving cysteine (Cy...

European Journal of Biochemistry, 1997

A new variant of cytochrome-c peroxidase in which the positively charged Arg48 present in the dis... more A new variant of cytochrome-c peroxidase in which the positively charged Arg48 present in the distal heme-binding pocket has been replaced with a Glu residue has been prepared and characterized to explore, in part, the possibility that a negative charge close to the heme could contribute to stabilization of a porphyrin-centered pi-cation radical in the compound I derivative of the variant. Between pH 4 and 8, this variant forms three pH-linked spectroscopic species. The electronic absorption and 1H-NMR spectra of the predominant form at low pH (HS1) are indicative of a high-spin, pentacoordinate heme iron system. Near neutral pH, a second high-spin species (HS2) is dominant, in which the heme iron center is hexacoordinated, with a water molecule as the sixth axial ligand. At high pH, the third form (LS) exhibits the spectroscopic characteristics of a low-spin, hexacoordinate heme center with bishistidine axial ligation. The apparent pKa values for these transitions are 4.4 and 7.4, respectively, in phosphate buffers and 5.0 and 7.1, respectively, in phosphate/nitrate buffers. Replacement of Arg48 with Glu reduces the thermal stability of the enzyme and also decreases the Fe(III)/Fe(II) reduction potential of the enzyme by approximately 50 mV relative to that of the wild-type enzyme. The stability of compound I formed by the variant is decreased although the rate at which it forms is just one order of magnitude less than that of the wild-type enzyme, thus confirming previous results which indicate that the function of residue 48 in the wild-type peroxidase is more related to the stability of compound I than to its formation [Erman, J. E., Vitello, L. B., Miller, M. A. & Kraut, J. (1992) J. Am. Chem. Soc. 114, 6592-6593; Vitello, L. B., Erman, J. E., Miller, M. A., Wang, J. & Kraut, J. (1993) Biochemistry 32, 9807-9818]. Stopped-flow studies failed to detect even transient formation of a porphyrin-centered radical following addition of hydrogen peroxide to the Fe(III)-enzyme. The consequences of this drastic electrostatic modification of the active site on the steady-state kinetics of the variant are relatively minor.

European Journal of Biochemistry, 1994

1H two-dimensional NMR spectroscopy has been applied to the oxidized form of cytochrome c 551 fro... more 1H two-dimensional NMR spectroscopy has been applied to the oxidized form of cytochrome c 551 from Rhodocyclus gelatinosus, which is paramagnetic with S = 1/2. The investigation has allowed a complete and unambiguous assignment of the heme protons and some residues around the heme. We have learned that: the conformation of the axial methionine is equal to that of horse heart cytochrome c and different from two isoenzymes of the same cytochrome c 551 from a different strain; pKa of 6.6 +/- 0.3 has been detected through the shift variations of seventh propionate protons. The detailed differences with other cytochromes c in the hyperfine shifts are discussed.

Supramolecular Chemistry, 2010

Journal of the American Chemical Society, 1999

The 800-MHz 1H NMR spectra of oxidized plastocyanin from spinach are here reported. All hyperfine... more The 800-MHz 1H NMR spectra of oxidized plastocyanin from spinach are here reported. All hyperfine-shifted signals have been assigned through saturation transfer with the reduced diamagnetic species. To detect the copper (II)-bound cysteine β-CH2 signals, a technique has been applied which is based on irradiation of regions where such signals are expected but not detected, and the corresponding saturation transfer on the reduced species is observed. At the end, a full spectrum is reconstructed which permits, for the first time, the ...

Journal of the American Chemical Society, 2002

Proton NMR spectra of FeIII-FeII recombinant single polypeptide human PAP (recHPAP) have been mea... more Proton NMR spectra of FeIII-FeII recombinant single polypeptide human PAP (recHPAP) have been measured at, above, and below its pH optimum, as have the spectra of inhibited forms containing fluoride and phosphate, analogues of the substrates hydroxide and phosphate esters, respectively. The results demonstrate that binding of inhibitory anions to the dinuclear mixed-valent site of recHPAP is controlled by protonation of a ligand to the dinuclear center. Thus, the group that is responsible for pKa,1 in the enzymatic activity versus pH profile functions as a &amp;quot;gatekeeper&amp;quot;, whose protonation state controls anion binding to the mixed-valent dinuclear site. The correlation between the pKa values observed in kinetics studies and for the spectroscopic changes strongly suggests that this group is the nucleophilic hydroxide that attacks the phosphate ester substrate.

Journal of the American Chemical Society, 2001

The NMR solution structure of oxidized plastocyanin from the cyanobacterium Synechocystis PCC6803... more The NMR solution structure of oxidized plastocyanin from the cyanobacterium Synechocystis PCC6803 is here reported. The protein contains paramagnetic copper(II), whose electronic relaxation times are quite unfavorable for NMR solution studies. The structure has been solved on the basis of 1041 meaningful NOESY cross-peaks, 18 1D NOEs, 26 T(1) values, 96 dihedral angle constraints, and 18 H-bonds. The detection of broad hyperfine-shifted signals and their full assignment allowed the identification of the copper(II) ligands and the determination of the Cu-S-C-H dihedral angle for the coordinated cysteine. The global root-mean-square deviation from the mean structure for the solution structure family is 0.72 +/- 0.14 and 1.16 +/- 0.17 A for backbone and heavy atoms, respectively. The structure is overall quite satisfactory and represents a breakthrough, in that it includes paramagnetic copper proteins among the metalloproteins for which solution structures can be afforded. The comparison with the available X-ray structure of a triple mutant is also performed.