RESONANCE RAMAN DETECTION OF 2 CONFORMERS FOR THE CYANIDE ADDUCT OF CYTOCHROME-C PEROXIDASE AND THEIR PH-DEPENDENCE

The resonance Raman spectra of the adducts of cytochrome c peroxidase with four cyanide isotopomers ((CN)-C-12-N-14-, (CN-)-C-13-N-14, (CN-) -C-12-N-15 and (CN-)-C-13-N-15) are presented. The spectra reveal the presence of two conformers, an essentially linear conformer whose v(Fe -C) and delta(FeCN) modes occur at 445 cm(-1) and 407 cm(-1), respecti vely (at pH 10) and a bent conformer which exhibits two modes at 355 c m(-1) and 445 cm(-1) (at pH 10). At pH values below 7, the linear form v(Fe-C) shifts to similar to 455 cm(-1) in response to protonation of the distal histidyl imidazole (which strengthens the Fe-C bond). The stretching mode of the bent form experiences only a slight shift. This smaller shift of the bent form is consistent with the proposal that t he bent form originates as a consequence of hydrogen bond formation wi th the off-axis proton doner group of arginine (which is protonated at both pH values). The overall spectral response to pH changes reflects interactions with the bound ligand which may be important for heterol ytic bond cleavage.