Decoupling of the bc1 complex in S. cerevisiae; point mutations affecting the cytochrome b gene bring new information about the structural aspect of the proton translocation.
Four mutations in the mitochondrial cytochrome b of S. cerevisiae have been characterized with respect to growth capacities, catalytic properties, ATP/2e- ratio, and transmembrane potential. The respiratory-deficient mutant G137E and the three pseudo-wild type revertants E137 + I147F, E137 + C133S, and E137 + N256K were described previously (Tron and Lemesle-Meunier, 1990; Di Rago et al., 1990a). The mutant G137E is unable to grow on respiratory substrates but its electron transfer activity is partly conserved and totally inhibited by antimycin A. The secondary mutations restore the respiratory growth at variable degree, with a phosphorylation efficiency of 12-42% as regards the parental wild type strain, and result in a slight increase in the various electron transfer activities at the level of the whole respiratory chain. The catalytic efficiency for ubiquinol was slightly (G137E) or not affected (E137 + I147F, E137 + C133S, and E137 + N256K) in these mutants. Mutation G137E induces a decrease in the ATP/2e- ratio (50% of the W.T. value) and transmembrane potential (60% of the W.T. value) at the bc1 level, whereas the energetic capacity of the cytochrome oxidase is conserved. Secondary mutations I147F, C133S, and N256K partly restore the ATP/2e- ratio and the transmembrane potential at the bc1 complex level. The results suggest that a partial decoupling of the bc1 complex is induced by the cytochrome b point mutation G137E. In the framework of the protonmotive Q cycle, this decoupling can be explained by the existence of a proton wire connecting centers P and N in the wild type bc1 complex which may be amplified or uncovered by the G137E mutation when the bc1 complex is functioning.
Bruel
C
C
Laboratoire de Bioénergétique et Ingénierie des Protéines, C.N.R.S., Marseille, France.
Manon
S
S
Guérin
M
M
Lemesle-Meunier
D
D
D000595:Amino Acid Sequence; D003573:Cytochrome b Group / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004579:Electron Transport; D014450:Electron Transport Complex III / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D007425:Intracellular Membranes / Q000502:physiology; D007700:Kinetics; D008564:Membrane Potentials; D008928:Mitochondria / Q000201:enzymology / Q000502:physiology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010101:Oxygen Consumption; D017354:Point Mutation; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*
D003573:Cytochrome b Group; D014450:Electron Transport Complex III