engleski

tRNA dependent amino acid recognition by yeast seryl-tRNA synthetase

The formation of aminoacyl-tRNA, catalyzed by aminoacyl-tRNA synthetases, is a crucial step in maintaining the fidelity of protein biosynthesis. Sequence-specific interactions between these enzymes and their cognate tRNAs both ensure accurate RNA recognition and prevent the binding of non-cognate substrates. An investigation of the structure/function relationship in Saccharomyces cerevisiae seryl-tRNA synthetase (SerRS) revealed that RNA:protein interactions also affect the amino acid affinity of the enzyme. Yeast cytoplasmic SerRS is a homodimeric class II aminoacyl-tRNA synthetase. It shares significant sequence homology with and has the same oligomeric structure as its prokaryotic counterparts, which were crystallized in other laboratories. This facilitated yeast SerRS modeling studies and the alignment-guided mutagenesis of its SES1 gene. We examined the effects of amino acid alterations in the active site and the truncation of the C-terminal non-catalytic domain (which characterizes only eukaryotic SerRS enzymes) on the substrate recognition. Steady-state kinetic analyses of the purified mutant SerRS proteins, in conjunction with in vivo complementation studies, suggest that yeast seryl-tRNA synthetase undergoes a conformational change induced by positioning the 3'end of tRNA into the active site. Even subtle perturbation of SerRS structure interfere with the proposed structural flexibility of the enzyme, which may lower the accuracy of aminoacylation. In order to determine whether tRNA-dependent amino acid recognition is a general property of serine system, we have recently cloned and overexpressed two organellar seryl-tRNA synthetases (one from yeast and the other from maize) whose catalytic mechanisms are currently under the study.

aminoacyl-tRNA synthetase; tRNA

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