Cytochrome rC 552, Formed during Expression of the Truncated, Thermus thermophilus Cytochrome c 552 Gene in the Cytoplasm of Escherichia coli, Reacts Spontaneously To Form Protein-Bound 2-Formyl-4-vinyl (Spirographis) Heme

Expression of the truncated (lacking an N-terminal signal sequence) structural gene of Thermus thermophilus cytochrome c 552 in the cytoplasm of Escherichia coli yields both dimeric (rC 557) and monomeric (rC 552) cytochrome c-like proteins [Keightley, J. A., et al. (1998) J. Biol. Chem. 273, 12006−12016], which form spontaneously without the involvement of cytochrome c maturation factors. Cytochrome rC 557 is comprised of a dimer and has been structurally characterized [McRee, D., et al. (2001) J. Biol. Chem. 276, 6537−6544]. Unexpectedly, the monomeric rC 552 transforms spontaneously to a cytochrome-like chromophore having, in its reduced state, the Q oo transition (α-band) at 572 nm (therefore called p572). The X-ray crystallographic structure of rC 552, at 1.41 Å resolution, shows that the 2-vinyl group of heme ring I is converted to a [heme-CO-CH2-S-CH2-Cα] conjugate with cysteine 11. Electron density maps obtained from isomorphous crystals of p572 at 1.61 Å resolution reveal that the 2-vinyl group has been oxidized to a formyl group. This explains the lower energy of the Q oo transition, the presence of a new, high-frequency band in the resonance Raman spectra at 1666 cm-1 for oxidized and at 1646 cm-1 for reduced samples, and the greatly altered, paramagnetically shifted 1H NMR spectrum observed for this species. The overall process defines a novel mechanism for oxidation of the 2-vinyl group to a 2-formyl group and adds to the surprising array of chemical reactions that occur in the interaction of heme with the CXXCH sequence motif in apocytochromes c.