Coq3 and Coq4 define a polypeptide complex in yeast mitochondria for the biosynthesis of coenzyme Q.
Coenzyme Q (Q) is a redox active lipid essential for aerobic respiration in eukaryotes. In Saccharomyces cerevisiae at least eight mitochondrial polypeptides, designated Coq1-Coq8, are required for Q biosynthesis. Here we present physical evidence for a coenzyme Q-biosynthetic polypeptide complex in isolated mitochondria. Separation of digitonin-solubilized mitochondrial extracts in one- and two-dimensional Blue Native PAGE analyses shows that Coq3 and Coq4 polypeptides co-migrate as high molecular mass complexes. Similarly, gel filtration chromatography shows that Coq1p, Coq3p, Coq4p, Coq5p, and Coq6p elute in fractions higher than expected for their respective subunit molecular masses. Coq3p, Coq4p, and Coq6p coelute with an apparent molecular mass exceeding 700 kDa. Coq3 O-methyltransferase activity, a surrogate for Q biosynthesis and complex activity, also elutes at this high molecular mass. We have determined the quinone content in lipid extracts of gel filtration fractions by liquid chromatography-tandem mass spectrometry and find that demethoxy-Q(6) is enriched in fractions with Coq3p. Co-precipitation of biotinylated-Coq3 and Coq4 polypeptide from digitonin-solubilized mitochondrial extracts shows their physical association. This study identifies Coq3p and Coq4p as defining members of a Q-biosynthetic Coq polypeptide complex.
Marbois
Beth
B
Department of Chemistry and Biochemistry and the Molecular Biology Institute, UCLA, Los Angeles, California 90095, USA.
Gin
Peter
P
Faull
Kym F
KF
Poon
Wayne W
WW
Lee
Peter T
PT
Strahan
Jeff
J
Shepherd
Jennifer N
JN
Clarke
Catherine F
CF
D016227:Benzoquinones / Q000032:analysis; D019921:Biotinylation; D055598:Chemical Phenomena; D011232:Chemical Precipitation; D002627:Chemistry, Physical; D002850:Chromatography, Gel; D002853:Chromatography, Liquid; D004072:Digitonin; D013058:Mass Spectrometry; D008780:Methyltransferases / Q000737:chemistry / Q000378:metabolism*; D008928:Mitochondria / Q000737:chemistry*; D024101:Mitochondrial Proteins; D008970:Molecular Weight; D012441:Saccharomyces cerevisiae / Q000648:ultrastructure*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D012995:Solubility; D014451:Ubiquinone / Q000096:biosynthesis* / Q000737:chemistry
D016227:Benzoquinones; C433869:COQ4 protein, S cerevisiae; D024101:Mitochondrial Proteins; D029701:Saccharomyces cerevisiae Proteins; C032165:ubiquinone 6; D014451:Ubiquinone; C004532:quinone; D008780:Methyltransferases; C070696:3,4-dihydroxy-5-hexaprenylbenzoate methyltransferase; D004072:Digitonin