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Purification, crystallization and preliminary X-ray analysis of the dissimilatory sulfite reductase from Desulfovibrio vulgaris Miyazaki F.

ABSTRACT: Dissimilatory sulfite reductase (Dsr) plays an important role in sulfate respiration in many sulfate-reducing bacteria. Dsr from Desulfovibrio vulgaris Miyazaki F has been purified and crystallized at 277?K using the sitting-drop vapour-diffusion method with PEG 3350 and potassium thiocyanate as precipitants. A data set was collected to 3.7?Å resolution from a single crystal at 100?K using synchrotron radiation. The Dsr crystal belonged to space group P4(1)2(1)2, with unit-cell parameters a = b = 163.26, c = 435.32?Å. The crystal structure of Dsr was determined by the molecular-replacement method based on the three-dimensional structure of Dsr from D. vulgaris Hildenborough. The crystal contained three ?(2)?(2)?(2) units per asymmetric unit, with a Matthews coefficient (V(M)) of 2.35?Å(3)?Da(-1); the solvent content was estimated to be 47.7%.

SUBMITTER: Ogata H

PROVIDER: S-EPMC3001650 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature

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Purification, crystallization and preliminary X-ray analysis of the dissimilatory sulfite reductase from Desulfovibrio vulgaris Miyazaki F.

Ogata Hideaki H Shomura Yasuhito Y Goenka Agrawal Aruna A Kaur Amrit Pal AP Gärtner Wolfgang W Higuchi Yoshiki Y Lubitz Wolfgang W

Acta crystallographica. Section F, Structural biology and crystallization communications 20101028 Pt 11

Dissimilatory sulfite reductase (Dsr) plays an important role in sulfate respiration in many sulfate-reducing bacteria. Dsr from Desulfovibrio vulgaris Miyazaki F has been purified and crystallized at 277 K using the sitting-drop vapour-diffusion method with PEG 3350 and potassium thiocyanate as precipitants. A data set was collected to 3.7 Å resolution from a single crystal at 100 K using synchrotron radiation. The Dsr crystal belonged to space group P4(1)2(1)2, with unit-cell parameters a = b ...[more]

PMID: 21045297

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Project description:The precise interpretation of environmental sulfur isotope records requires a quantitative understanding of the biochemical controls on sulfur isotope fractionation by the principle isotope-fractionating process within the S cycle, microbial sulfate reduction (MSR). Here we provide the only direct observation of the major ((34)S/(32)S) and minor ((33)S/(32)S, (36)S/(32)S) sulfur isotope fractionations imparted by a central enzyme in the energy metabolism of sulfate reducers, dissimilatory sulfite reductase (DsrAB). Results from in vitro sulfite reduction experiments allow us to calculate the in vitro DsrAB isotope effect in (34)S/(32)S (hereafter, [Formula: see text]) to be 15.3 ± 2‰, 2?. The accompanying minor isotope effect in (33)S, described as [Formula: see text], is calculated to be 0.5150 ± 0.0012, 2?. These observations facilitate a rigorous evaluation of the isotopic fractionation associated with the dissimilatory MSR pathway, as well as of the environmental variables that govern the overall magnitude of fractionation by natural communities of sulfate reducers. The isotope effect induced by DsrAB upon sulfite reduction is a factor of 0.3-0.6 times prior indirect estimates, which have ranged from 25 to 53‰ in (34)?DsrAB. The minor isotope fractionation observed from DsrAB is consistent with a kinetic or equilibrium effect. Our in vitro constraints on the magnitude of [Formula: see text] is similar to the median value of experimental observations compiled from all known published work, where (34)? r-p = 16.1‰ (r-p indicates reactant vs. product, n = 648). This value closely matches those of MSR operating at high sulfate reduction rates in both laboratory chemostat experiments ([Formula: see text] 17.3 ± 1.5‰, 2?) and in modern marine sediments ([Formula: see text] 17.3 ± 3.8‰). Targeting the direct isotopic consequences of a specific enzymatic processes is a fundamental step toward a biochemical foundation for reinterpreting the biogeochemical and geobiological sulfur isotope records in modern and ancient environments.

Dataset Information

Purification, crystallization and preliminary X-ray analysis of the dissimilatory sulfite reductase from Desulfovibrio vulgaris Miyazaki F.

Publications

Purification, crystallization and preliminary X-ray analysis of the dissimilatory sulfite reductase from Desulfovibrio vulgaris Miyazaki F.

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