c-type cytochrome assembly in Saccharomyces cerevisiae: a key residue for apocytochrome c1/lyase interaction.
The electron transport chains in the membranes of bacteria and organelles generate proton-motive force essential for ATP production. The c-type cytochromes, defined by the covalent attachment of heme to a CXXCH motif, are key electron carriers in these energy-transducing membranes. In mitochondria, cytochromes c and c(1) are assembled by the cytochrome c heme lyases (CCHL and CC(1)HL) and by Cyc2p, a putative redox protein. A cytochrome c(1) mutant with a CAPCH heme-binding site instead of the wild-type CAACH is strictly dependent upon Cyc2p for assembly. In this context, we found that overexpression of CC(1)HL, as well as mutations of the proline in the CAPCH site to H, L, S, or T residues, can bypass the absence of Cyc2p. The P mutation was postulated to shift the CXXCH motif to an oxidized form, which must be reduced in a Cyc2p-dependent reaction before heme ligation. However, measurement of the redox midpoint potential of apocytochrome c(1) indicates that neither the P nor the T residues impact the thermodynamic propensity of the CXXCH motif to occur in a disulfide vs. dithiol form. We show instead that the identity of the second intervening residue in the CXXCH motif is key in determining the CCHL-dependent vs. CC(1)HL-dependent assembly of holocytochrome c(1). We also provide evidence that Cyc2p is dedicated to the CCHL pathway and is not required for the CC(1)HL-dependent assembly of cytochrome c(1).
Corvest
Vincent
V
Department of Chemistry and Biochemistry, Center for Biotechnology and Genomics, Texas Tech University, Lubbock, TX 79409, USA.
Murrey
Darren A
DA
Bernard
Delphine G
DG
Knaff
David B
DB
Guiard
Bernard
B
Hamel
Patrice P
PP
D001665:Binding Sites; D002352:Carrier Proteins / Q000378:metabolism*; D045304:Cytochromes c / Q000096:biosynthesis / Q000378:metabolism*; D004579:Electron Transport; D004591:Electrophoresis, Polyacrylamide Gel; D015966:Gene Expression Regulation, Fungal; D006418:Heme / Q000378:metabolism; D008190:Lyases / Q000378:metabolism*; D008928:Mitochondria / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000378:metabolism*; D009154:Mutation; D010084:Oxidation-Reduction; D016133:Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*
C083336:CYC2 protein, S cerevisiae; D002352:Carrier Proteins; D024101:Mitochondrial Proteins; D029701:Saccharomyces cerevisiae Proteins; D006418:Heme; D045304:Cytochromes c; D008190:Lyases; C031581:cytochrome C synthetase