Cytochrome b558/566 from the archaeon Sulfolobus acidocaldarius. A novel highly glycosylated, membrane-bound b-type hemoprotein.
In this study we re-examined the inducible cytochrome b558/566 from the archaeon Sulfolobus acidocaldarius (DSM 639), formerly thought to be a component of a terminal oxidase (Becker, M., and Schäfer, 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 b558/566 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 alpha-helical membrane anchors flanking the majority of a mainly beta-pleated sheet structure containing unusually high amounts 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 b558/566 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.
Hettmann
T
T
Institut für Biochemie, Medizinische Universität zu Lübeck, Ratzeburger Allee 160, 23538 Lübeck, Germany.
Schmidt
C L
CL
Anemüller
S
S
Zähringer
U
U
Moll
H
H
Petersen
A
A
Schäfer
G
G
D000595:Amino Acid Sequence; D001483:Base Sequence; D002849:Chromatography, Gas; D003573:Cytochrome b Group / Q000378:metabolism*; D004269:DNA, Bacterial; D006031:Glycosylation; D006420:Hemeproteins / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D019255:NADPH Oxidases; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D016945:Sulfolobus acidocaldarius / Q000201:enzymology*
D003573:Cytochrome b Group; D004269:DNA, Bacterial; D006420:Hemeproteins; D008565:Membrane Proteins; C020730:cytochrome b566; C036888:cytochrome b558; D019255:NADPH Oxidases