A CRYO-CRYSTALLOGRAPHIC REDOX STUDY OF THE STRUCTURAL CHANGES CAUSED BY MUTATION OF KEY AMINO ACIDS AT THE FMN BINDING SITE OF A FLAVODOXIN M.A. Walsh,^1,2 A. McCarthy,² T. Higgins,³ P. O'Farrell³ and S.G. Mayhew.^{3. 1}European Molecular Biology Laboratory, Hamburg, Germany. ²Department of Chemistry, University College, Galway, Ireland ³Department of Biochemistry, University College, Dublin, Ireland

The flavodoxins are a group of small flavoproteins which function as electron carriers in low potential redox reactions. They contain a single molecule of non-covalently bound flavin mononucleotide, (FMN) as their sole prosthetic group. The FMN cofactor can exist in three oxidation states: oxidized, semiquinone and hydroquinone. When bound to protein, the redox potentials of FMN for both the oxidized/semiquinone, (E2) and semiquinone/hydroquione, (E1) are significantly shifted. These shifts in redox potential, are essential for flavodoxin to carry out its physiological role. The interactions between the protein and FMN which cause the large shifts in redox potential are not yet fully understood. A number of site directed mutagenesis studies have therefore been carried out to asses the role of specific amino acids at the FMN binding site of flavodoxin.^1-3 We have carried out an extensive crystallographic study of a number of the site mutations in their differing oxidation states at the FMN binding site The results show the FMN binding site to be more flexible than previously envisioned and further structural studies should allow a more clearer understanding of the protein/FMN functioning

1. Curley GP, Carr MC, O'Farrell PA, Mayhew, SG Voordouw, G (1991). Flavins and Flavoproteins, 1990, Berlin: Walter de Gruyter,429

2. Swenson, RP. & Krey, GD. (1994) Biochemistry 33, 8805

3. Swenson, RP & Zhou, Z. (1995) Biochemistry 34, 3183