title: Biochemical and Spectroscopic Studies of Epoxyqueuosine Reductase: A Novel Iron-Sulfur Cluster- and Cobalamin-Containing Protein Involved in the Biosynthesis of Queuosine.
abstract: Queuosine is a hypermodified nucleoside present in the wobble position of tRNAs with a 5'-GUN-3' sequence in their anticodon (His, Asp, Asn, and Tyr). The 7-deazapurine core of the base is synthesized de novo in prokaryotes from guanosine 5'-triphosphate in a series of eight sequential enzymatic transformations, the final three occurring on tRNA. Epoxyqueuosine reductase (QueG) catalyzes the final step in the pathway, which entails the two-electron reduction of epoxyqueuosine to form queuosine. Biochemical analyses reveal that this enzyme requires cobalamin and two [4Fe-4S] clusters for catalysis. Spectroscopic studies show that the cobalamin appears to bind in a base-off conformation, whereby the dimethylbenzimidazole moiety of the cofactor is removed from the coordination sphere of the cobalt but not replaced by an imidazole side chain, which is a hallmark of many cobalamin-dependent enzymes. The bioinformatically identified residues are shown to have a role in modulating the primary coordination sphere of cobalamin. These studies provide the first demonstration of the cofactor requirements for QueG.
authors.affiliation: †Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona 85721, United States.
authors.forename: Zachary D
authors.initials: ZD
authors.lastname: Miles
authors.affiliation: ‡Department of Chemistry, University of California-Davis, Davis, California 95616, United States.
authors.forename: William K
authors.initials: WK
authors.lastname: Myers
authors.affiliation: †Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona 85721, United States.
authors.forename: William M
authors.initials: WM
authors.lastname: Kincannon
authors.affiliation: ‡Department of Chemistry, University of California-Davis, Davis, California 95616, United States.
authors.forename: R David
authors.initials: RD
authors.lastname: Britt
authors.affiliation: †Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona 85721, United States.
authors.forename: Vahe
authors.initials: V
authors.lastname: Bandarian
mesh_terms: D001412:Bacillus subtilis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002384:Catalysis; D007506:Iron-Sulfur Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009704:Nucleoside Q / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014805:Vitamin B 12 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D007506:Iron-Sulfur Proteins; D009704:Nucleoside Q; D010088:Oxidoreductases; D014805:Vitamin B 12