title: Utility of muropeptide ligase for identification of inhibitors of the cell wall biosynthesis enzyme MurF.
abstract: MurF is a key enzyme in the biosynthesis of the bacterial cell wall in both gram-positive and gram-negative bacteria. This enzyme has not been extensively exploited as a drug target, possibly due to the difficulty in obtaining one of the substrates, UDP-MurNAc-L-Ala-gamma-D-Glu-meso-diaminopimelate, which is usually purified from bacteria. We have identified putative inhibitors of Escherichia coli MurF by a binding assay, thus bypassing the need for substrate. Inhibition of enzymatic activity was demonstrated in a high-performance liquid chromatography-based secondary assay with UDP-MurNAc-L-Ala-gamma-D-Glu-diaminopimelate substrate prepared in a novel way by using muropeptide ligase enzyme to add UDP-MurNAc to synthetic L-Ala-gamma-D-Glu-diaminopimelate; the substrate specificity of muropeptide ligase for peptides containing L-Lys in place of diaminopimelate was also investigated. Using the muropeptide ligase-generated MurF substrate, a thiazolylaminopyrimidine series of MurF enzyme inhibitors with 50% inhibitory concentration values as low as 2.5 microM was identified.
authors.affiliation: Johnson & Johnson Pharmaceutical Research & Development, LLC, 1000 Route 202, Raritan, New Jersey 08869, USA. ebaum@prdus.jnj.com
authors.forename: Ellen Z
authors.initials: EZ
authors.lastname: Baum
authors.forename: Steven M
authors.initials: SM
authors.lastname: Crespo-Carbone
authors.forename: Darren
authors.initials: D
authors.lastname: Abbanat
authors.forename: Barbara
authors.initials: B
authors.lastname: Foleno
authors.forename: Amy
authors.initials: A
authors.lastname: Maden
authors.forename: Raul
authors.initials: R
authors.lastname: Goldschmidt
authors.forename: Karen
authors.initials: K
authors.lastname: Bush
mesh_terms: D002473:Cell Wall / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000302:isolation & purification* / Q000494:pharmacology; D004926:Escherichia coli / Q000737:chemistry / Q000201:enzymology*; D010453:Peptide Synthases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010457:Peptidoglycan / Q000096:biosynthesis*
chemical_list: D004791:Enzyme Inhibitors; D010457:Peptidoglycan; D010453:Peptide Synthases; C060628:UDP-N-acetylmuramoylalanyl-D-glutamyllysine-D-alanyl-D-alanine ligase; C049145:UDP-N-acetylmuramoylalanyl-D-glutamate-2,6-diaminopimelate ligase