Assignment of the histidine axial ligands to the cytochrome bH and cytochrome bL components of the bc1 complex from Rhodobacter sphaeroides by site-directed mutagenesis.
The cytochrome b subunit of the bc1 complex contains two cytochrome components, cytochrome bH and cytochrome bL. Sequence comparisons of this polypeptide from a number of organisms have revealed four invariant histidines which have been postulated to be the heme ligands for the two protoheme IX prosthetic groups. In Rhodobacter sphaeroides, these correspond to His97, His111, His198, and His212. In this paper, the results of amino acid substitutions at each of these positions are reported. Replacement of His97 by either Asp or Asn and of His198 by Asn or Tyr resulted in loss of both cytochrome components. However, His111Asn, His111Asp, and His212Asp all resulted in the selective loss of cytochrome bH and the retention of cytochrome bL. Furthermore, flash kinetics studies show that the myxothiazol-sensitive quinol oxidase (Qz) site associated with cytochrome bL is still functional. These data support the assignment of the axial ligands to cytochrome bH (His111 and His212) and cytochrome bL (His97 and His198). This pairing is consistent with current models of the cytochrome b subunit with eight transmembrane alpha-helices.
Yun
C H
CH
Department of Biochemistry and Chemistry, University of Illinois, Urbana 61801.
Crofts
A R
AR
Gennis
R B
RB
D000595:Amino Acid Sequence; D001483:Base Sequence; D015153:Blotting, Western; D003062:Codon; D003573:Cytochrome b Group / Q000737:chemistry* / Q000235:genetics; D014450:Electron Transport Complex III / Q000737:chemistry* / Q000235:genetics; D006418:Heme / Q000378:metabolism; D006639:Histidine / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008024:Ligands; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010084:Oxidation-Reduction; D012242:Rhodobacter sphaeroides / Q000201:enzymology* / Q000235:genetics
D003062:Codon; D003573:Cytochrome b Group; D008024:Ligands; D006418:Heme; D006639:Histidine; D014450:Electron Transport Complex III