title: Identification of the binding site of the quinone-head group in mitochondrial Coq10 by photoaffinity labeling.
abstract: Mitochondrial Coq10 is a ubiquinone (UQ)-binding protein that is a member of the steroidogenic acute regulatory protein (StAR)-related lipid transfer (START) domain superfamily. Deletion of the COQ10 gene was previously shown to cause a marked respiratory defect in Saccharomyces cerevisiae and Schizosaccharomyces pombe, which indicated that Coq10 may support efficient electron transfer between the respiratory complexes; however, its physiological role remains elusive. To elucidate the role of Coq10, we attempted to identify the binding site of UQ in recombinant S. pombe Coq10 expressed in an Escherichia coli cell membrane through photoaffinity labeling with the photoreactive UQ probe, UQ-1, in combination with biotinylation of the labeled peptide by means of the so-called click chemistry. Comprehensive proteomic analyses revealed that the quinone-head ring of UQ-1 specifically binds to the N-terminal region of Phe39–Lys45 of Coq10, which corresponds to the ligand-binding pocket of many proteins containing the START domain. The labeling was completely suppressed in the presence of an excess amount of artificial short-chain UQ analogues, such as UQ2. In the Phe39Ala and Pro41Ala mutants, the extents of labeling were ∼40 and ∼60%, respectively, of that of wild-type Coq10. While Coq10 has been thought to bind UQ, our work first provides the direct evidence of Coq10 accommodating the quinone-head ring of UQ in its START domain. On the basis of these results, the physiological role of Coq10 has been discussed.
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Murai
authors.forename: Kohei
authors.initials: K
authors.lastname: Matsunobu
authors.forename: Sawako
authors.initials: S
authors.lastname: Kudo
authors.forename: Kentaro
authors.initials: K
authors.lastname: Ifuku
authors.forename: Makoto
authors.initials: M
authors.lastname: Kawamukai
authors.forename: Hideto
authors.initials: H
authors.lastname: Miyoshi
mesh_terms: D001665:Binding Sites / Q000235:genetics; D004926:Escherichia coli / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D010750:Phosphoproteins / Q000737:chemistry; D019920:Photoaffinity Labels / Q000737:chemistry; D012568:Schizosaccharomyces / Q000378:metabolism; D014451:Ubiquinone / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics; D000098682:Steroidogenic Acute Regulatory Protein
chemical_list: D010750:Phosphoproteins; D019920:Photoaffinity Labels; D000098682:Steroidogenic Acute Regulatory Protein; D014451:Ubiquinone; C024989:coenzyme Q10