title: Stop codon recognition in the early-diverged protozoans Giardia lamblia and Trichomonas vaginalis.
abstract: Two classes of polypeptide release factors (RFs) are responsible for maintaining accuracy in translation termination; however, their detailed mechanism of action and evolutionary history of these factors remain elusive. The structure and function of RFs vary in bacteria and eukaryotes, a fact that is suggestive of evolutionary changes in the translation termination system. Giardia lamblia (Diplomonada) and Trichomonas vaginalis (Parabasalia) are considered as early-diverged eukaryotes. The class II release factor, eRF3, of Giardia (Gl-eRF3) appears to have only one domain that corresponds to EF-1α and lacks the N-terminal domain, similar to that of eRF3 of other organisms. In the present study, we show that the chimeric molecules Gl/Sc eRF1 and Tv/Sc eRF1, which are composed of the N-terminal domain of Gl-eRF1 or Tv-eRF1, fused to the core domain (M and C domain) of Saccharomyces cerevisiae eRF1 (Sc-eRF1), resulting in loss of the RF properties of the N-terminal domain. This suggests that the conformation of eRF1 for stop codon recognition in Giardia and Trichomonas varies from the eRF1s of other eukaryotes, including ciliates and yeast. Further studies using intra-N-terminal chimeras of eRF1 indicated that the combination of the GTS loop and NIKS motif from Gl-eRF1 and the Y-C-F motif from Sc-eRF1within the N terminal domain of hybrid eRF1 could restore UGA, but not UAG and UGA recognition. In contrast, the combination of the GTS loop and the NIKS motif of Sc-eRF1 and the Y-C-F motif of Gl-eRF1 could restore UAG and UAA recognition, but not UGA recognition. Thus, these results confirm the findings of previous studies that three motifs in eRF1 are necessary for discrimination of the three bases of stop codons. The NIKS motif is responsible for recognition of the first two bases of UAA and UAG, and the Y-C-F motif identifies the second base of UGA by Gl-eRF1. Amino acid residue substitutions in Gl/Sc-eRF1 by corresponding residues of Sc-eRF1 could change and even restore RF activity, further suggesting different conformation of eRF1 are used for stop codon recognition in Giardia and in Saccharomyces.
authors: Key Laboratory of Chemical Biology and Molecular Engineering, Ministry of Education, Institute of Biotechnology, Shanxi University, Taiyuan 030006, China. Electronic address: bfchai@sxu.edu.cn.
authors: Baofeng
authors: B
authors: Chai
authors: Faculty of Environment and Economics, Shanxi University of Finance and Economics, Taiyuan 030006, China.
authors: Cui
authors: C
authors: Li
authors: Key Laboratory of Chemical Biology and Molecular Engineering, Ministry of Education, Institute of Biotechnology, Shanxi University, Taiyuan 030006, China.
authors: Jingfei
authors: J
authors: Yu
authors: Key Laboratory of Chemical Biology and Molecular Engineering, Ministry of Education, Institute of Biotechnology, Shanxi University, Taiyuan 030006, China.
authors: Yanrong
authors: Y
authors: Hao
authors: Key Laboratory of Chemical Biology and Molecular Engineering, Ministry of Education, Institute of Biotechnology, Shanxi University, Taiyuan 030006, China.
authors: Ping
authors: P
authors: Guo
authors: Key Laboratory of Chemical Biology and Molecular Engineering, Ministry of Education, Institute of Biotechnology, Shanxi University, Taiyuan 030006, China.
authors: Quan
authors: Q
authors: Shen
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D018388:Codon, Terminator; D018503:Genome, Protozoan; D016829:Giardia lamblia / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D010454:Peptide Termination Factors / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics; D014246:Trichomonas vaginalis / Q000235:genetics*
chemical_list: D018388:Codon, Terminator; D010454:Peptide Termination Factors
keywords: eRF1; evolution; protist; stop codon recognition