Journal
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Volume 349, Issue 4, Pages 1250-1257Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.bbrc.2006.08.166
Keywords
Escherichia coli; essential genes; drug targets; antibacterial compounds; over-expression; triclosan; phosphomycin
Categories
Funding
- NIGMS NIH HHS [S06 GM008101, R25 GM061331, S06GM008101] Funding Source: Medline
- NIMHD NIH HHS [P20MD001824, P20 MD001824] Funding Source: Medline
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With the advancement of high throughput screening, it has become easier and faster to discover hit compounds that inhibit proliferation of bacterial cells. However, development in technologies used to identify cellular targets of potent antibacterial inhibitors has lagged behind. Here, we describe a novel strategy of target identification for antibacterial inhibitors using an array of Escherichia coli clones each over-expressing one essential protein. In a proof-of-concept study, eight essential genes were cloned into pLex5BA vector under the control of an inducible promoter. Over-expression of target proteins was confirmed. For two clones, one over-expressing FabI and the other over-expressing MurA enzymes, the host cells became 17- and 139-fold more resistant to the specific inhibitors triclosan and phosphomycin, respectively, while the susceptibility of other clones towards these inhibitors remained unchanged after induction of gene expression. Target identification via target protein over-expression was demonstrated using both mixed clone and individual clone assay formats. (c) 2006 Elsevier Inc. All rights reserved.
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