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

Jorge Caldeira, P. Nuno Palma, Manuela Regalla, Jorge Lampreia, Juan Calvete, Wolfram Schäfer, Jean Legall, Isabel MOURA, José J. G. MOURA

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Abstract

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.

Original languageEnglish
Pages (from-to)987-995
Number of pages9
JournalEuropean Journal Of Biochemistry
Volume220
Issue number3
DOIs
Publication statusPublished - 1 Jan 1994

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