The Structure of the Oxidized Form of Clostridial Flavodoxin at 1.9-A Resolution

Abstract An electron density map of the oxidized form of flavodoxin from Clostridium strain MP has been calculated to a resolution of 1.9 A employing isomorphous and anomalous scattering data from two heavy atom derivatives, Smiii and Aui. In the region from 3.0 to 1.9 A, the largest structure amplitudes (85% of the total | F | s) have been included in the computation. The over-all figure of merit is 0.808. The molecular model resulting from interpretation of this map has a chain length of 138 residues, in accord with the sequence (Tanaka, M., Haniu, M., Yasunobu, K. T., and Mayhew, S. G. (1974) J. Biol. Chem. 249, 4393–4396). As reported earlier, the molecule possesses a high degree of secondary structure, including a central parallel sheet and four helices. More than 70% of the residues appear to participate in backbone hydrogen bonding. Atomic coordinates, incorporating the sequence data, have been optimized by use of Diamond's model-building program. The flavin mononucleotide prosthetic group adopts a somewhat extended conformation. As expected for the oxidized state, the isoalloxazine ring is planar. It is partly shielded from solvent by a tryptophan residue and interacts on its inner surface with methionine. Protein atoms are also in appropriate positions for interaction with atoms N-1, O-2, and N-3 of the flavin ring, but N-5 appears too far removed to form a hydrogen bond with the apoprotein. The ribityl phosphate moiety also makes multiple contacts with protein groups. Two of the three O' oxygens are in positions where they may participate in hydrogen bonding; the phosphate is closely surrounded by four hydroxyamino acids and four backbone NH groups. However, the prosthetic group is accessible on one side to solvent. Peaks likely to represent solvent can be observed near the dimethylbenzene end of the isoalloxazine ring and adjacent to the 3'-ribityl oxygen. While the mutual arrangement of protein and ribityl phosphate is nearly identical with that found by Watenpaugh et al. (Watenpaugh, K. D., Sieker, L. C., Jensen, L. H., LeGall, J., and Dubourdieu, M. (1972) Proc. Nat. Acad. Sci. U. S. A. 69, 3185–3188) in Desulfovibrio vulgaris flavodoxin, the orientation and interactions of the flavin ring differ strikingly in the two flavodoxins. The structure of the oxidized form of Clostridium MP flavodoxin is very similar to that deduced earlier for the semiquinone form at 3.25 A, despite large differences in the x-ray intensities (Ri = 0.66).