Cytochrome c' of Methylococcus capsulatus Bath.

Abstract

Cytochrome c' was isolated from the obligate methylotroph Methylococcus capsulatus Bath. The native and subunit molecular masses of the cytochrome were 34.9 kDa and 16.2 kDa, respectively, with an isoelectric pH of 7.0. The amino acid composition and N-terminal amino acid sequence were consistent with identification of the protein as a cytochrome c'. The electron paramagnetic resonance spectrum of the monoheme cytochrome indicated the presence of a high spin, S = 5/2, heme center that is diagnostic of cytochromes c'. The optical absorption spectra of ferric or ferrous cytochrome c' were also characteristic of cytochromes c'. The ferrocytochrome bound carbon monoxide and nitric oxide, but not isocyanide, cyanide, or azide. Changes in physical properties due to binding of CO or NO to some other c'-type cytochromes have been interpreted as an indication of dimer dissociation. In the case of cytochrome c' from M. capsulatus Bath, analytical ultracentrifugation of the ferricytochrome, the ferrocytochrome, and the ferrocytochrome-CO complex indicate that the changes induced by binding of CO are conformational and are not consistent with dimer dissociation. EPR spectra show that cytochrome c' was reduced in the presence of hydroxylamine only when in a complex with cytochrome P-460. The value of the midpoint potential, Em 7.0, was -250 mV for cytochrome c' from M. capsulatus Bath, which is well below the range of values reported for other cytochromes c'. The values of midpoint potentials for cytochrome P-460 (Em 7.0 = -300 mV to -380 mV) and cytochrome C555 (Em 7.0 = +175 mV to +195 mV) are less than and greater than, respectively, the value for cytochrome c' and suggest the possibility that the latter may function as an electron shuttle between cytochrome P-460 and cytochrome C555.