Effect of the ionic environment on the molecular structure of bacteriophage SPP1 portal protein

Bacteriophage SPP1 portal protein is a large cyclical homo-oligomer composed of 13 subunits. The solution structure and assembly behavior of this protein with high-point rotational symmetry was characterized. The purified protein was present as a monodisperse population of 13-mers, named gp6H, at univalent salt concentrations in the hundred millimolar range (>= 250 m m NaCl) or in the presence of bivalent cations in the millimolar range (>= 5 m m MgCl2). Gp6H had a slightly higher sedimentation coefficient, a smaller shape-dependent frictional ratio, and a higher rate of intersubunit cross-linking in the presence of magnesium than in its absence. In the absence of bivalent cations and at univalent salt concentrations below 250 m m, the 13-mer molecules dissociated partially into stable monomers, named gp6L. The monomer had a somewhat different shape from the subunit present in the 13-mer, but maintained a defined tertiary structure. The association-dissociation equilibrium was mainly between the monomer and the 13-mer with a minor population of intermediate oligomers. Their interconversion was strongly influenced by the ionic environment. Under physiological conditions, the concentration of Mg2+ found in the Bacillus subtilis cytoplasm (10-50 m m) probably promotes complete association of gp6 into 13-mer rings with a compact conformation

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.publishersversionpublishe