Bacterial Prolyl-tRNA Synthetase

So this article talks about a specific feature of bacterial prolyl-tRNA synthetase. The article explains how the substrate specificity of bacterial prolyl-tRNA synthetase's editing domain is controlled by a tunable hydrophobic pocket. Aminoacyl-tRNA synthetases are catalysts that covalently attach amino acids onto their respective tRNA's. High accuracy in this reaction is essential to the proper decoding of genetic information for if a charged tRNA molecule contains the wrong amino acid, the incorrect amino acid will be added to the polypeptide chain, thus making a faulty protein. One way the cell tries to prevent this mishap is by proofreading newly synthesized aminoacyl-tRNA molecules. Prolyl-tRNA synthetase (ProRS) mischarges tRNA(Pro) with alanine or cysteine because of their smaller or similar sizes relative to cognate proline. Mischarged Ala-tRNA(Pro) is hydrolyzed by the editing domain, INS, present in most bacterial ProRSs. On the other hand, the INS domain is unable to deacylate Cys-tRNA(Pro). Cys-tRNA(Pro) is hydrolyzed exclusively by a free-standing trans editing domain known as YbaK.

Researchers used computational and experimental approaches to probe the molecular basis of INS domain alanine specificity. In Escherichia coli ProRS, the methyl side chain of alanine binds in a well-defined hydrophobic pocket characterized by conserved residues I263, L266, and K279 and partially conserved residue T277. Site-specific mutation of these residues leads to a significant loss in Ala-tRNA(Pro) hydrolysis, and altering the size of the pocket modulates the substrate specificity. Surprisingly, one ProRS INS domain variant displayed a complete switch in substrate specificity from alanine to cysteine.