Primary sequence, oxidation‐reduction potentials and tertiary‐structure prediction of Desulfovibrio desulfuricans ATCC 27774 flavodoxin

Flavodoxin was isolated and purified from Desulfovibrio desulfuricans ATCC 27774, a sulfate‐reducing organism that can also utilize nitrate as an alternative electron acceptor. Mid‐point oxidation‐reduction potentials of this flavodoxin were determined by ultraviolet/visible and EPR methods coupled to potentiometric measurements and their pH dependence studied in detail. The redox potential E₂, for the couple oxidized/semiquinone forms at pH 6.7 and 25°C is –40 mV, while the value for the semiquinone/hydroquinone forms (E₁), at the same pH, –387 mV. E₂ varies linearly with pH, while E₁ is independent of pH at high values. However, at low pH (< 7.0), this value is less negative, compatible with a redox‐linked protonation of the flavodoxin hydroquinone. A comparative study is presented for Desulfovibrio salexigens NCIB 8403 flavodoxin [Moura, I., Moura, J. J. G., Bruschi, M. & LeGall, J. (1980) Biochim. Biophys. Acta 591, 1–8]. The complete primary amino acid sequence was obtained by automated Edman degradation from peptides obtained by chemical and enzymic procedures. The amino acid sequence was confirmed by FAB/MS. Using the previously determined tridimensional structure of Desulfovibrio vulgaris flavodoxin as a model [similarity, 48.6%; Watenpaugh, K. D., Sieker, L. C., Jensen, L. H., LeGall, J. & Dubourdieu M. (1972) Proc. Natl Acad. Sci. USA 69, 3185–3188], the tridimensional structure of D. desulfuricans ATCC 27774 flavodoxin was predicted using AMBER force‐field calculations.