5ucm
Crystal Structure of Prolyl-tRNA Synthetase from Pseudomonas aeruginosaCrystal Structure of Prolyl-tRNA Synthetase from Pseudomonas aeruginosa
Structural highlights
FunctionSYP_PSEAE Catalyzes the attachment of proline to tRNA(Pro) in a two-step reaction: proline is first activated by ATP to form Pro-AMP and then transferred to the acceptor end of tRNA(Pro). As ProRS can inadvertently accommodate and process non-cognate amino acids such as alanine and cysteine, to avoid such errors it has two additional distinct editing activities against alanine. One activity is designated as 'pretransfer' editing and involves the tRNA(Pro)-independent hydrolysis of activated Ala-AMP. The other activity is designated 'posttransfer' editing and involves deacylation of mischarged Ala-tRNA(Pro). The misacylated Cys-tRNA(Pro) is not edited by ProRS. Publication Abstract from PubMedPseudomonas aeruginosa is an opportunistic multi-drug resistant pathogen implicated as a causative agent in nosocomial and community acquired bacterial infections. The gene encoding prolyl-tRNA synthetase (ProRS) from P. aeruginosa was overexpressed in Escherichia coli and the resulting protein was characterized. ProRS was kinetically evaluated and the KM values for interactions with ATP, proline, and tRNA were 154, 122, and 5.5 muM, respectively. The turn-over numbers, kcat (obs) , for interactions with these substrates were calculated to be 5.5, 6.3, and 0.2 s(-1) , respectively. The crystal structure of the alpha2 form of P. aeruginosa ProRS was solved to 2.60 A resolution. The amino acid sequence and X-ray crystal structure of P. aeruginosa ProRS was analyzed and compared with homologs in which the crystal structures have been solved. The amino acids that interact with ATP and proline are well conserved in the active site region and overlay of the crystal structure with ProRS homologs conforms to a similar overall three-dimensional structure. ProRS was developed into a screening platform using scintillation proximity assay (SPA) technology and used to screen 890 chemical compounds, resulting in the identification of two inhibitory compounds, BT06A02 and BT07H05. This work confirms the utility of a screening system based on the functionality of ProRS from P. aeruginosa. Characterization and structure determination of prolyl-tRNA synthetase from Pseudomonas aeruginosa and development as a screening platform.,Pena N, Dranow DM, Hu Y, Escamilla Y, Bullard JM Protein Sci. 2019 Jan 21. doi: 10.1002/pro.3579. PMID:30666738[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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