Aminoacyl-tRNA substrate and enzyme backbone atoms contribute to translational quality control by YbaK.
JOURNAL OF PHYSICAL CHEMISTRY B(2013)
摘要
Amino acids are covalently attached to their corresponding transfer RNAs (tRNAs) by aminoacyl-tRNA synthetases. Proofreading mechanisms exist to ensure that high fidelity is maintained in this key step in protein synthesis. Prolyl-tRNA synthetase (ProRS) can misacylate cognate tRNA(Pro) with Ala and Cys. The cis-editing domain of ProRS (INS) hydrolyzes Ala-tRNA(Pro), whereas Cys-tRNA(Pro) is hydrolyzed by a single domain editing protein, YbaK, in trans. Previous studies have proposed a model of substrate-binding by bacterial YbaK and elucidated a substrate-assisted mechanism of catalysis. However, the microscopic steps in this mechanism have not been investigated In this work, we carried out biochemical experiments together with a detailed hybrid quantum mechanics/molecular mechanics study to investigate the mechanism of catalysis by Escherichia coli YbaK The results support a mechanism wherein cyclization of the substrate Cys results in cleavage of the Cys-tRNA ester bond Protein side chains do not play a significant role in YbaK catalysis. Instead, protein backbone atoms play crucial roles in stabilizing the transition state, while the product is stabilized by the 2'-OH of the tRNA.
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关键词
carrier proteins,hydrolysis,escherichia coli,biocatalysis,quantum theory,protein biosynthesis
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