Cytochrome b(558/566) from the Archaeon Sulfolobus acidocaldarius. A novel highly glycosylated, membrane-bound B-type hemoprotein

In this study we re-examined the inducible cytochrome b(558/566) from the archaeon Sulfolobus acidocaldarius (DSM 639), formerly thought to be a component of a terminal oxidase (Becker, M., and Schafer, G. (1991) FEBS Lett, 291, 331-335). An improved purification method increased the yield of the protein and allowed more detailed investigations. Its molecular mass and heme content have been found to be 64,210 Da and 1 mol of heme/mol of protein, respectively. It is only detectable in cells grown at low oxygen tensions. The composition of the growth medium also exerts significant influence on the cytochrome b(558/566) content of S. acidocaldarius membranes. The cytochrome exhibits an extremely high redox potential of +400 mV and shows no CO reactivity; a ligation other than a His/His-coordination of axial ligands appears likely. It turned out to be highly glycosylated (more than 20% of its molecular mass are sugar residues) and is probably exposed to the outer surface of the plasma membrane. The sugar moiety consists of several O-glycosidically linked mannoses and at least one N- glycosidically linked hexasaccharide comprising two glucoses, two mannoses, and two N-acetyl-glucosamines. The gene of the cytochrome (cbsA) has been sequenced, revealing an interesting predicted secondary structure with two putative α-helical membrane anchors flanking the majority of a mainly β- pleated sheet structure containing unusually high mounts of serine and threonine. A second gene (cbsB) was found to be cotranscribed. The latter displays extreme hydrophobicity and is thought to form a functional unit with cytochrome b(558/566) in vivo, although it did not copurify with the latter. Sequence comparisons show no similarity to any entry in data banks indicating that this cytochrome is indeed a novel kind of b-type hemoprotein. A cytochrome c analogous function in the pseudoperiplasmic space of S. acidocaldarius is discussed.