Q-band electron nuclear double resonance (ENDOR) and X-band EPR of the sulfobetaine 12 heat-treated cytochrome c oxidase complex.
Heat treatment of the bovine cytochrome c oxidase complex in the zwitterionic detergent sulfobetaine 12 (SB-12) results in loss of subunit III and the appearance of a type II copper center as characterized by electron paramagnetic resonance (EPR) spectroscopy. Previous authors (Nilsson, T., Copeland, R. A., Smith, P. A., and Chan, S. I. (1988) Biochemistry 27, 8254-8260) have interpreted this type II copper center as a modified version of the CuA site. By using electron nuclear double resonance spectroscopy, it is found that the CuA proton and nitrogen resonances remain present in the SB-12 heat-treated enzyme and that three new nitrogen resonances appear having hyperfine coupling constants consistent with histidine ligation. These hyperfine coupling constants correlate well with those recently found for the CuB histidines from the cytochrome aa(3)-600 quinol oxidase from Bacillus subtilis (Fann, Y. C., Ahmed, I., Blackburn, N. J., Boswell, J. S., Verkhovskaya, M. L., Hoffman, B. M., and Wikstr9om, M. (1995) Biochemistry 34, 10245-10255). In addition, the total EPR-detectable copper concentration per enzyme molecule approximately doubles upon SB-12 heat treatment. Finally, the observed type II copper EPR spectrum is virtually indistinguishable from the EPR spectrum of CuB of the as-isolated cytochrome bo3 complex from Escherichia coli. These data indicate that the type II copper species that appears results from a breaking of the strong antiferromagnetic coupling of the heme a3-CuB binuclear center.
Musser
S M
SM
Division of Chemistry and Chemical Engineering, Arthur Amos Noyes Laboratory of Chemical Physics, California Institute of Technology, Pasadena 91125, USA.
Fann
Y C
YC
Gurbiel
R J
RJ
Hoffman
B M
BM
Chan
S I
SI
D000818:Animals; D002417:Cattle; D003300:Copper / Q000737:chemistry; D003902:Detergents; D004578:Electron Spin Resonance Spectroscopy; D003576:Electron Transport Complex IV / Q000737:chemistry*; D006358:Hot Temperature; D000644:Quaternary Ammonium Compounds / Q000737:chemistry; D013056:Spectrophotometry, Ultraviolet
D003902:Detergents; D000644:Quaternary Ammonium Compounds; C034730:zwittergent 3-12; D003300:Copper; D003576:Electron Transport Complex IV