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Study of genes from Xenopus laevis and Caenorhabditis elegans homologues to Saccharomyces cerevisiae SUP35, controlling ambiguity of translation in yeast

It was proposed, based on genetic data, that both yeast Sup45 and Sup35 proteins might be involved in the maintenance of translation fidelity or/and in translation termination. The Saccharomyces cerevisiae SUP35 gene has been identified and sequenced. Yeast Sup35p is composed of at least two domains: the amino (N) and the carboxy (C) terminal domains (253 and 432 amino acids, respectively). The C-domain exhibits GTP binding motifs. The C-domain of the Sup35-like proteins is conserved even between distant species like yeast and human while the N-terminal parts of these proteins are entirely different. We have now identified a Xenopus laevis SUP35 gene using yeast and human probes. The deduced amino acid sequence of the C-domain of the X. Laevis Sup 35-like protein (designated as Sup 35Cp) is similar to that of human GSPT1p, S. Cerevisiae and Pichia pinus Sup35p indicating the high conservation of the primary structure of this domain. All these proteins exhibit GTP binding motifs (G1-G4) in the C-domain similar to those in the other G proteins, in particular in elongation factor EF-1&;alpha and prokaryotic RF3, that have to be responsible for guanine nucleotide binding and GTP hydrolysis. Plasmids carrying XSUP35C could compensate temperature-sensitivity but much less efficiently in comparison with plasmids carrying GSPT1 gene or yeast SUP35 gene. They could not complement respiratory sensitivity, also they do not influence the suppression of nonsense mutations and they are unable to compensate inviability caused by disruption of SUP35 gene. This result could be explained by a functional difference of the yeast and Xenopus Sup35 proteins caused by the differences in their primary structure, or by the low level of the XSUP35 expression in yeast. Since it has been shown earlier that yeast Sup45 protein (Sup45p) belongs to eRF1 protein family it seems reasonable to assume that Sup35 protein (Sup35p) could be a candidate for the putative non-identified eRF. In common work with L. Kisselev laboratory it was shown that the newly sequenced GTP binding Sup35-like protein from Xenopus laevis, termed eRF3, exhibits in vitro three important functional properties: (i) although being inactive as eRF on its own, it greatly stimulates eRF1 activity in the presence of GTP and low concentrations of stop codons, resembling the properties of prokaryotic RF3, (ii) it binds and probably hydrolyses GTP, and (iii) it binds to the eRF1.

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Université de Rennes I
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