Editor-in-Chief: Brüne, Bernhard
Editorial Board Member: Buchner, Johannes / Lei, Ming / Ludwig, Stephan / Sies, Helmut / Turk, Boris / Wittinghofer, Alfred
SCImago Journal Rank (SJR) 2015: 1.607
Source Normalized Impact per Paper (SNIP) 2015: 0.751
Impact per Publication (IPP) 2015: 2.609
Characterization of peptidyl-tRNA hydrolase encoded by open reading frame Rv1014c of Mycobacterium tuberculosis H37Rv
1Molecular and Structural Biology, Central Drug Research Institute, Lucknow 226 001, India
The first two authors contributed equally to this work.
2Molecular and Structural Biology, Central Drug Research Institute, Lucknow 226 001, India
3Molecular and Structural Biology, Central Drug Research Institute, Lucknow 226 001, India
4Molecular and Structural Biology, Central Drug Research Institute, Lucknow 226 001, India
Citation Information: Biological Chemistry. Volume 388, Issue 5, Pages 467–479, ISSN (Online) 14316730, ISSN (Print) 14374315, DOI: 10.1515/BC.2007.057, May 2007
- Published Online:
The enzyme peptidyl-tRNA hydrolase (Pth, EC 188.8.131.52) is essential for the viability of bacteria. The ORF Rv1014c of Mycobacterium tuberculosis H37Rv, designated as the mtpth gene, was cloned and over-expressed and the product was purified. Generation of polyclonal antibodies against the purified recombinant protein, termed MtPth, facilitated detection of endogenously expressed MtPth in M. tuberculosis H37Rv cell lysate. MtPth could release diacetyl-[3H]-lysine from diacetyl-[3H]-lysyl-tRNALys with Michaelis-Menten kinetic parameters of K m=0.7±0.2 μM and k cat=1.22±0.2 s-1. Transformation of a pTrc99c/mtpth vector allowed the growth of E. coli thermosensitive Pth(ts) mutant strain AA7852 at the non-permissive temperature of 42°C, demonstrating the in vivo activity of MtPth. In addition, at 39°C, over-expression of MtPth in AA7852 cells allowed the cells to remain viable in the presence of up to 200 μg/ml erythromycin. A 3D fold based on NMR and a structural model based on the E. coli Pth crystal structure were generated for MtPth. The essential nature of conserved active-site residues N12, H22 and D95 of MtPth for catalysis was demonstrated by mutagenesis and complementation in E. coli mutant strain AA7852. Thermal and urea/guanidinium chloride (GdmCl)-induced unfolding curves for MtPth indicate a simple two-state unfolding process without any intermediates.
Here you can find all Crossref-listed publications in which this article is cited. If you would like to receive automatic email messages as soon as this article is cited in other publications, simply activate the “Citation Alert” on the top of this page.