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T Petrova,
E Y Bezsudnova,
K M Boyko,
A V Mardanov,
K M Polyakov,
V V Volkov,
M Kozin,
N V Ravin,
I G Shabalin,
K G Skryabin, T N Stekhanova,
M V Kovalchuk,
V O Popov
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ABSTRACT: DNA ligases join single-strand breaks in double-stranded DNA by catalyzing the formation of a phosphodiester bond between adjacent 5'-phosphate and 3'-hydroxyl termini. Their function is essential for maintaining genome integrity in the replication, recombination and repair of DNA. High flexibility is important for the function of DNA ligase molecules. Two types of overall conformations of archaeal DNA ligase that depend on the relative position of the OB-fold domain have previously been revealed: closed and open extended conformations. The structure of ATP-dependent DNA ligase from Thermococcus sp. 1519 (LigTh1519) in the crystalline state determined at a resolution of 3.02 Å shows a new relative arrangement of the OB-fold domain which is intermediate between the positions of this domain in the closed and the open extended conformations of previously determined archaeal DNA ligases. However, small-angle X-ray scattering (SAXS) measurements indicate that in solution the LigTh1519 molecule adopts either an open extended conformation or both an intermediate and an open extended conformation with the open extended conformation being dominant.
Acta Crystallographica Section F Structural Biology and Crystallization Communications 12/2012; 68(Pt 12):1440-7. · 0.51 Impact Factor
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A V Lyashenko,
E Y Bezsudnova,
V M Gumerov,
A A Lashkov,
A V Mardanov,
A M Mikhailov,
K M Polyakov,
V O Popov,
N V Ravin,
K G Skryabin,
V K Zabolotniy, T N Stekhanova,
M V Kovalchuk
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ABSTRACT: Alcohol dehydrogenases belong to the oxidoreductase family and play an important role in a broad range of physiological processes. They catalyze the cofactor-dependent reversible oxidation of alcohols to the corresponding aldehydes or ketones. The NADP-dependent short-chain alcohol dehydrogenase TsAdh319 from the thermophilic archaeon Thermococcus sibiricus was overexpressed, purified and crystallized. Crystals were obtained using the hanging-drop vapour-diffusion method using 25%(w/v) polyethylene glycol 3350 pH 7.5 as precipitant. The crystals diffracted to 1.68 A resolution and belonged to space group I222, with unit-cell parameters a = 55.63, b = 83.25, c = 120.75 A.
Acta Crystallographica Section F Structural Biology and Crystallization Communications 06/2010; 66(Pt 6):655-7. · 0.51 Impact Factor
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ABSTRACT: DNA ligases catalyze the sealing of 5'-phosphate and 3'-hydroxyl termini at single-strand breaks in double-stranded DNA and their function is essential to maintain the integrity of the genome in DNA metabolism. An ATP-dependent DNA ligase from the archaeon Thermococcus sp. 1519 was overexpressed, purified and crystallized. Crystals were obtained using the hanging-drop vapour-diffusion method employing 35%(v/v) Tacsimate pH 7.0 as a precipitant and diffracted X-rays to 3.09 A resolution. They belonged to space group P4(1)2(1)2, with unit-cell parameters a = b = 79.7, c = 182.6 A.
Acta Crystallographica Section F Structural Biology and Crystallization Communications 05/2009; 65(Pt 4):368-71. · 0.51 Impact Factor
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ABSTRACT: Formate dehydrogenase (FDH) from the methylotrophic bacterium Pseudomonas sp. 101 catalyzes oxidation of formate to NI2 with the coupled reduction of nicotinamide adenine dinucleotide (NAD+). The three-dimensional structures of the apo form (the free enzyme) and the holo form (the ternary FDH-NAD+-azide complex) of FDH have been established earlier. In the present study, the structures of FDH complexes with formate are
solved at 2.19 and 2.28 Å resolution by the molecular replacement method and refined to the R factors of 22.3 and 20.5%, respectively. Both crystal structures contain four protein molecules per asymmetric unit. These
molecules form two dimers identical to the dimer of the apo form of FDH. Two possible formatebinding sites are found in the
active site of the FDH structure. In the complexes the sulfur atom of residue Cys354 exists in the oxidized state.
Crystallography Reports 06/2006; 51(4):627-631. · 0.47 Impact Factor
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M V Telkov,
G R Demina,
S A Voloshin,
E G Salina,
T V Dudik, T N Stekhanova,
G V Mukamolova,
K A Kazaryan,
A V Goncharenko,
M Young,
A S Kaprelyants
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ABSTRACT: The secreted Micrococcus luteus protein, Rpf, is required for successful resuscitation of dormant "non-culturable" M. luteus cells and for growth stimulation in poor media. The biochemical mechanism of Rpf action remained unknown. Theoretical predictions of Rpf domain architecture and organization, together with a recent NMR analysis of the protein structure, indicate that the conserved Rpf domain has a lysozyme-like fold. In the present study, we found that both the secreted native protein and the recombinant protein lyse crude preparations of M. luteus cell walls. They also hydrolyze 4-methylumbelliferyl-beta-D-N,N',N''-triacetylchitotrioside, a synthetic substrate for peptidoglycan muramidases, with optimum activity at pH 6. The Rpf protein also has weak proteolytic activity against N-CBZ-Gly-Gly-Arg-beta-naphthylamide, a substrate for trypsin-like enzymes. Rpf activity towards 4-methylumbelliferyl-beta-D-N,N',N''-triacetylchitotrioside was reduced when the glutamate residue at position 54, invariant for all Rpf family proteins and presumably involved in catalysis, was altered. The same amino acid substitution resulted in impaired resuscitation activity of Rpf. The data indicate that Rpf is a peptidoglycan-hydrolyzing enzyme, and strongly suggest that this specific activity is responsible for its growth promotion and resuscitation activity. A possible mechanism of Rpf-mediated resuscitation is discussed.
Biochemistry (Moscow) 05/2006; 71(4):414-22. · 1.06 Impact Factor
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ABSTRACT: Formate dehydrogenase (FDG) from methylotrophic bacteria Pseudomonas sp. 101 catalyzes the reaction of oxidation of the formate ion to carbon dioxide, which is accompanied by the reduction of nicotinamid
adenine dinucleotide (NAD+). The structures of the apo and holo (enzyme-NAD-azide triple complex) forms of the enzyme were determined earlier. In an
attempt to prepare a complex of FDG with the product of the enzymatic reaction (NADH), a new crystal modification of FDG is
obtained (space group P42212, a = b = 93.3 Å, c = 103.05 Å). The FDG structure is solved by the molecular replacement method and refined to R = 20.7%. The asymmetric part of the unit cell contains one FDG molecule. In contrast to the previously studied FDG structures,
the biologically active dimer is formed by the crystallographic rotation axis. A comparative structural analysis of the studied
form with the apo and holo forms of the enzyme is performed. The influence of the molecular structure on the environment in
the crystal is investigated.
Crystallography Reports 08/2005; 50(5):796-800. · 0.47 Impact Factor
