Great Britain Electron - Paramagnetic - Resonance Studies on Cobalt ( ll ) Carbonic Anhydrase LOW - SPIN CYANIDE COMPLEXES By STEPHEN

The low-spin cyanide complexes of three Co(ll) carbonic anhydrases were investigated by electron paramagnetic resonance (e.p.r.) at 9 and 35GHz. Well-defined and closely axial spectra were obtained only in the absence of oxygen. Several mole equivalents of cyanidewererequired for complete formation ofthecomplexes in frozen solution, although large excesses caused abstraction of the cobalt. Experiments with [13C]cyanide showed that the low-spin complexes contained two CNgroups in an environment similar to that ofthe in-plane ligands in [Co(CN)5s]3. A combined e.p.r. and spectrophotometric titration confirmed the presence oftwo CNligands. A 5-co-ordinate square pyramidal structure involving three protein ligands was proposed. The dicyanide complex could be oxygenated reversibly, producing a characteristic new e.p.r. spectrum. The 02 molecule was thought to occupy the remaining octahedral metal site in a formally Co(III) species. The optical spectrum of the dicyanide lacked the prominent d-d bands of the high-spin monocyanide. Both e.p.r. and optical data indicated that the low-spin complex was formed much more fully in frozen solution than at room temperature. Differences in behaviour between the highand low-activity enzymes suggested some variation in conformational flexibility at the metal binding site.