title: Relationship between the GTPase, metal-binding, and dimerization activities of E. coli HypB.
abstract: Biosynthesis of the metallocenter in the active site of the [NiFe] hydrogenase enzyme requires the accessory protein HypB, which is a metal-binding GTPase. In this study, the interplay between the individual activities of Escherichia coli HypB was examined. The full-length protein undergoes nucleotide-responsive dimerization that is disrupted upon mutation of L242 and L246 to alanine. This mutant HypB is monomeric under all of the conditions investigated but the inability of L242A/L246A HypB to dimerize does not abolish its GTPase activity and the monomeric protein has metal-binding behavior similar to that of wild-type HypB. Furthermore, expression of L242A/L246A HypB in vivo results in hydrogenase activity that is approximately half of the activity produced by the wild-type control, suggesting that dimerization of HypB does not have a critical role in the hydrogenase maturation pathway. In contrast, the GTPase activity of HypB is modulated by metal loading of the protein. These results provide insight into the role of HypB in hydrogenase biosynthesis.
authors.affiliation: Department of Chemistry, University of Toronto, 80 St. George Street, Toronto, ON, M5S 3H6, Canada.
authors.forename: Fang
authors.initials: F
authors.lastname: Cai
authors.forename: Thanh T
authors.initials: TT
authors.lastname: Ngu
authors.forename: Harini
authors.initials: H
authors.lastname: Kaluarachchi
authors.forename: Deborah B
authors.initials: DB
authors.lastname: Zamble
mesh_terms: D000818:Animals; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000378:metabolism; D008670:Metals / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization
chemical_list: D029968:Escherichia coli Proteins; C490324:HypB protein, E coli; D008670:Metals; D006153:Guanosine Diphosphate; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins