title: Uncoupling of initiation factor eIF5B/IF2 GTPase and translational activities by mutations that lower ribosome affinity.
abstract: Translation initiation factor eIF5B/IF2 is a GTPase that promotes ribosomal subunit joining. We show that eIF5B mutations in Switch I, an element conserved in all GTP binding domains, impair GTP hydrolysis and general translation but not eIF5B subunit joining function. Intragenic suppressors of the Switch I mutation restore general translation, but not eIF5B GTPase activity. These suppressor mutations reduce the ribosome affinity of eIF5B and increase AUG skipping/leaky scanning. The uncoupling of translation and eIF5B GTPase activity suggests a regulatory rather than mechanical function for eIF5B GTP hydrolysis in translation initiation. The translational defect suggests eIF5B stabilizes Met-tRNA(i)(Met) binding and that GTP hydrolysis by eIF5B is a checkpoint monitoring 80S ribosome assembly in the final step of translation initiation.
authors.affiliation: Laboratory of Gene Regulation and Development, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Byung-Sik
authors.initials: BS
authors.lastname: Shin
authors.forename: David
authors.initials: D
authors.lastname: Maag
authors.forename: Antonina
authors.initials: A
authors.lastname: Roll-Mecak
authors.forename: M Shamsul
authors.initials: MS
authors.lastname: Arefin
authors.forename: Stephen K
authors.initials: SK
authors.lastname: Burley
authors.forename: Jon R
authors.initials: JR
authors.lastname: Lorsch
authors.forename: Thomas E
authors.initials: TE
authors.lastname: Dever
mesh_terms: D000483:Alleles; D000595:Amino Acid Sequence / Q000235:genetics; D002455:Cell Division / Q000235:genetics; D005057:Eukaryotic Cells / Q000201:enzymology*; D020558:GTP Phosphohydrolases / Q000235:genetics* / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics; D015966:Gene Expression Regulation, Fungal / Q000235:genetics; D006160:Guanosine Triphosphate / Q000235:genetics / Q000378:metabolism; D009154:Mutation / Q000235:genetics*; D039881:Prokaryotic Initiation Factor-2 / Q000235:genetics* / Q000378:metabolism; D014176:Protein Biosynthesis / Q000235:genetics*; D012343:RNA, Transfer / Q000235:genetics / Q000378:metabolism; D012270:Ribosomes / Q000201:enzymology* / Q000235:genetics; D013489:Suppression, Genetic / Q000235:genetics; D013912:Threonine / Q000235:genetics / Q000378:metabolism; D015003:Yeasts
chemical_list: D039881:Prokaryotic Initiation Factor-2; D013912:Threonine; D006160:Guanosine Triphosphate; D012343:RNA, Transfer; D020558:GTP Phosphohydrolases