Examination of \u3cem\u3eKlebsiella pneumoniae\u3c/em\u3e 5\u27-Methylthioadenosine/S-Adenosylhomocysteine Nucleosidase and 5-Methylthioribose Kinase

Klebsiella pneumoniae is an opportunistic bacterial pathogen that is emerging as a major global threat as an infectious agent. This organism, along with many other pathogens, possesses a broad suite of antibiotic resistances that can make treatment exceedingly difficult. As such, the impetus for creating novel antibiotics is at an all-time high despite the fact that pharmaceutical investment into drug development is at an all-time low. S-adenosylmethionine (SAM) is a universally utilized metabolite involved in a wide array of biosynthetic processes, and whose products are catabolized by a different set of enzymes in mammals than used by many bacteria, protozoa, and plants. This divergence provides two specific targets for enzymatic inhibition in K. pneumoniae: 5’-Methylthioadenosine/S-adenosylhomocysteine Nucleosidase (MTN) and 5-Methylthioribose Kinase (MTRK). In order to examine the potential effects of drugs targeting these enzymes, recombinant proteins from K. pneumoniae were isolated and purified, and a profile of their respective kinetic activities were determined. The MTRK enzyme was identified as using an ordered sequential mechanism, and shows preferential binding towards substrate analogs with hydrophobic 5-alkylthio substitutions. In addition, binding affinities of transition state-analogs and novel non-nucleoside small molecule inhibitors of MTN were quantified. Finally, the effects of MTN inhibitors on K. pneumoniae whole-cell growth, and gene expression were assessed. A number of the inhibitors demonstrated strong affinities for the MTN enzyme and induced alterations in cellular growth, autoinducer response, and the expression of genes associated with virulence