THE 2.6-ANGSTROM REFINED STRUCTURE OF THE ESCHERICHIA-COLI RECOMBINANT SACCHAROMYCES-CEREVISIAE FLAVOCYTOCHROME B(2) SULFITE COMPLEX

Flavocytochrome b(2) from Saccharomyces cerevisiae catalyzes the oxida tion of L-lactate to pyruvate and the electron transfer to cytochrome c in the mitochondrial intermembrane space. It is a homotetramer with a molecular weight of 4 x 58 kDa, each monomer of which is composed of 2 distinct domains, the one carrying FMN and the other, a ''b(5)-like '' heme. The native structure has been described at a resolution of 2. 4 Angstrom (Xia ZX, Mathews FS, 1990, J Mol Biol 212:837-863). The hem e domains protrude from the central body of the tetramer consisting of the 4 FMN binding domains. Because only 2 heme domains are visible in the electron density map, the other 2 are probably disordered. We cry stallized the Escherichia coli recombinant flavocytochrome b(2) from S . cerevisiae inhibited by sulfite. Although the crystals were obtained under very different conditions from those of the pyruvate-containing native enzyme, they were found to be isostructural (P 3(2) 2 1, a = b = 164.5 Angstrom, c = 114.0 Angstrom). The 2.6-Angstrom X-ray structu re was extensively refined with X-PLOR (R = 17.3%), which made it poss ible to describe in detail the recombinant flavocytochrome b(2) molecu lar structure. There exist few differences between the native and reco mbinant structures, in line with the fact that they show similar kinet ic behavior, and they further confirm the intrinsic mobility of the he me domain (Labeyrie F, Beloil JC, Thomas MA, 1988, Biochim Biophys Act a 953:134-141). This structure will be used as a starting model in the structural resolution of flavocytochrome b(2) point mutants.