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Publication Abstract from PubMed

The tRNA (m(1)G37) methyltransferase TrmD catalyzes m(1)G formation at position 37 in many tRNA isoacceptors and is essential in most bacteria, which positions it as a target for antibiotic development. In spite of its crucial role, little is known about TrmD in Pseudomonas aeruginosa (PaTrmD), an important human pathogen. Here we present detailed structural, substrate and kinetic properties of PaTrmD. Mass spectrometric analysis confirmed the G36G37-containing tRNAs Leu(GAG), Leu(CAG), Leu(UAG), Pro(GGG), Pro(UGG), Pro(CGG), and His(GUG) as PaTrmD substrates. Analysis of steady-state kinetics with S-adenosyl-L-methionine (SAM) and tRNA(Leu(GAG)) showed that PaTrmD catalyzes the two-substrate reaction by way of a ternary-complex, while isothermal titration calorimetry revealed that SAM and tRNA(Leu(GAG)) bind to PaTrmD independently, each with a dissociation constant of 14 +/- 3 microM. Inhibition by the SAM analog sinefungin was competitive with respect to SAM (Ki = 0.41 +/- 0.07 microM) and uncompetitive for tRNA (Ki = 6.4 +/- 0.8 microM). A set of crystal structures of the homodimeric PaTrmD protein bound to SAM and sinefungin provide the molecular basis for enzyme competitive inhibition and identify the location of the bound divalent ion. These results provide insights into PaTrmD as a potential target for the development of antibiotics.

Crystal structure and catalytic mechanism of the essential m(1)G37 tRNA methyltransferase TrmD from Pseudomonas aeruginosa.,Jaroensuk J, Wong YH, Zhong W, Liew CW, Maenpuen S, Sahili AE, Atichartpongkul S, Chionh YH, Nah Q, Thongdee N, McBee ME, Prestwich EG, DeMott MS, Chaiyen P, Mongkolsuk S, Dedon P, Lescar J, Fuangthong M RNA. 2019 Aug 9. pii: rna.066746.118. doi: 10.1261/rna.066746.118. PMID:31399541^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.