Identification, sequence determination, and expression of the flavodoxin gene from Desulfovibrio salexigens

Helms, Larry R.; Krey, Grigorios; Swenson, Richard P.

doi:10.1016/0006-291x(90)92393-e

Cited by 28 publications

(12 citation statements)

References 10 publications

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“…Propagation of conformational changes induced by mutations at position 98 to the adjacent 60's loop in the flavin binding site is not totally unexpected. Preliminary molecular modeling studies of several flavodoxins from the Desulfovibrio family suggest that sequence differences noted in the 90's loop can affect the conformation of residues in the 60's loop (Helms et al, 1990;Helms & Swenson, 1991;Helms and Swenson, unpublished results). The presence of a methionine residue at position 62 in the Desulfovibrio desulfuricans (Essex 6) flavodoxin, a position generally occupied by an acidic residue in this family, seems to influence the conformation of tyrosine-98 and the adjacent region (Helms & Swenson, 1992).…”

Section: Discussionmentioning

confidence: 97%

Structural changes caused by site-directed mutagenesis of tyrosine-98 in Desulfovibrio vulgaris flavodoxin delineated by 1H and 15N NMR spectroscopy: implications for redox potential modulation

Stockman

Richardson²,

Swenson³

1994

Biochemistry

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Flavodoxins mediate electron transfer at low redox potential between the prosthetic groups of other proteins. Interactions between the protein and the flavin mononucleotide cofactor shift both the oxidized/semiquinone and semiquinone/hydroquinone redox potentials significantly from their free-in-solution values. In order to investigate the possible role that the tyrosine at position 98 plays in this process, we have used heteronuclear three-dimensional NMR spectroscopy to determine the solution conformation of wild-type and four position-98 mutants, Y98W, Y98H, Y98A, and Y98R, of Desulfovibrio vulgaris flavodoxin. Assigned 1H and 15N resonances indicate that the secondary structure and topology of the proteins are identical. However, residues that undergo substantial mutation-induced changes in chemical shift are spread throughout the flavin cofactor binding site. Distance and dihedral angle constraints were used to generate solution structures for the wild-type and mutant proteins. Collectively, the mutant proteins have no gross conformational changes in the flavin binding site. The changes that do occur are minor and result from the different packing interactions required to accommodate the new side chain at position-98. The solvent accessibility and electrostatic nature of the flavin binding site in the mutant proteins are compared to those of the wild-type structure. The structural data support the hypothesis that the very low midpoint of the semiquinone/hydroquinone couple in the wild-type protein is modulated to a large extent by the energetically unfavorable formation of the flavin hydroquinone anion in the apolar environment of the flavin binding site.

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Section: Discussionmentioning

confidence: 97%

Structural changes caused by site-directed mutagenesis of tyrosine-98 in Desulfovibrio vulgaris flavodoxin delineated by 1H and 15N NMR spectroscopy: implications for redox potential modulation

Stockman

Richardson²,

Swenson³

1994

Biochemistry

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“…PCC 7119 and nonphotosynthetic bacterium Desulfovibrio salexigens were successfully overproduced in Escherichia coli (17,18). The rFlvd of Synechococcus sp.…”

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confidence: 99%

Measurement of Photosystem I Activity with Photoreduction of Recombinant Flavodoxin

Zhao

Bryant

1998

Analytical Biochemistry

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“…Our general strategy for identification of the reductase gene was patterned after that previously described (Krey et al, 1988;Helms et al, 1990):…”

Section: Resultsmentioning

confidence: 99%

“…The transformation mixture was then plated onto GeneScreen Plus membranes (DuPont) that had been placed on LB/amp plates. All plates were then replica plated onto Genescreen Plus (duplicate copies were made for each plate), and colonies were allowed to grow at 37 OC to a diameter of 1-2 nm for the colony hybridizations described below (Helms et al, 1990).…”

Section: Methodsmentioning

confidence: 99%

Primary structure of the assimilatory-type sulfite reductase from Desulfovibrio vulgaris (Hildenborough): cloning and nucleotide sequence of the reductase gene

Tan

Helms²,

Swenson³

et al. 1991

Biochemistry

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The nucleotide sequence encoding the structural gene (651 bp) and flanking regions for the assimilatory-type sulfite reductase from the sulfate-reducing bacterium Desulfovibrio vulgaris (Hildenborough) was determined after cloning a 1.4 kb HindIII/SalI genomic fragment possessing the gene into Bluescript pBS(+)KS. The primary structure of the protein was deduced, and the molecular mass of the apoprotein was estimated as 24 kDa. The amino acid sequence of the polypeptide shows some similarities at putative [Fe4S4] cluster binding sites in comparison with the heme protein subunit of the larger Escherichia coli and Salmonella typhimurium sulfite reductases and spinach nitrite reductase. This is the first reported sequence of a member of a new class of low molecular weight assimilatory sulfite-reducing enzymes recently identified in a number of anaerobic bacteria [Moura, I., Lina, A. R., Moura, J. J. G., Xavier, A. V., Fauque, G., Peck, H. D., & Le Gall, J. (1986) Biochem. Biophys. Res. Commun. 141, 1032-1041].

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Identification, sequence determination, and expression of the flavodoxin gene from Desulfovibrio salexigens

Cited by 28 publications

References 10 publications

Structural changes caused by site-directed mutagenesis of tyrosine-98 in Desulfovibrio vulgaris flavodoxin delineated by 1H and 15N NMR spectroscopy: implications for redox potential modulation

Structural changes caused by site-directed mutagenesis of tyrosine-98 in Desulfovibrio vulgaris flavodoxin delineated by 1H and 15N NMR spectroscopy: implications for redox potential modulation

Measurement of Photosystem I Activity with Photoreduction of Recombinant Flavodoxin

Primary structure of the assimilatory-type sulfite reductase from Desulfovibrio vulgaris (Hildenborough): cloning and nucleotide sequence of the reductase gene

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