title: Mutations in the highly conserved GGQ motif of class 1 polypeptide release factors abolish ability of human eRF1 to trigger peptidyl-tRNA hydrolysis.
abstract: Although the primary structures of class 1 polypeptide release factors (RF1 and RF2 in prokaryotes, eRF1 in eukaryotes) are known, the molecular basis by which they function in translational termination remains obscure. Because all class 1 RFs promote a stop-codon-dependent and ribosome-dependent hydrolysis of peptidyl-tRNAs, one may anticipate that this common function relies on a common structural motif(s). We have compared amino acid sequences of the available class 1 RFs and found a novel, common, unique, and strictly conserved GGQ motif that should be in a loop (coil) conformation as deduced by programs predicting protein secondary structure. Site-directed mutagenesis of the human eRF1 as a representative of class 1 RFs shows that substitution of both glycyl residues in this motif, G183 and G184, causes complete inactivation of the protein as a release factor toward all three stop codons, whereas two adjacent amino acid residues, G181 and R182, are functionally nonessential. Inactive human eRF1 mutants compete in release assays with wild-type eRF1 and strongly inhibit their release activity. Mutations of the glycyl residues in this motif do not affect another function, the ability of eRF1 together with the ribosome to induce GTPase activity of human eRF3, a class 2 RF. We assume that the novel highly conserved GGQ motif is implicated directly or indirectly in the activity of class 1 RFs in translation termination.
authors.affiliation: Engelhardt Institute of Molecular Biology, Moscow, Russia.
authors.forename: L Y
authors.initials: LY
authors.lastname: Frolova
authors.forename: R Y
authors.initials: RY
authors.lastname: Tsivkovskii
authors.forename: G F
authors.initials: GF
authors.lastname: Sivolobova
authors.forename: N Y
authors.initials: NY
authors.lastname: Oparina
authors.forename: O I
authors.initials: OI
authors.lastname: Serpinsky
authors.forename: V M
authors.initials: VM
authors.lastname: Blinov
authors.forename: S I
authors.initials: SI
authors.lastname: Tatkov
authors.forename: L L
authors.initials: LL
authors.lastname: Kisselev
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D017124:Conserved Sequence; D020558:GTP Phosphohydrolases / Q000378:metabolism; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010443:Peptide Chain Termination, Translational; D010454:Peptide Termination Factors / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; C489002:ETF1 protein, human; D010454:Peptide Termination Factors; D012346:RNA, Transfer, Amino Acyl; D015534:Trans-Activators; C019037:tRNA, peptidyl-; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases