Affinity Selection-Mass Spectrometry Identifies a Novel Antibacterial RNA Polymerase Inhibitor.
The growing prevalence of drug resistant bacteria is a significant global threat to human health. The antibacterial drug rifampin, which functions by inhibiting bacterial RNA polymerase (RNAP), is an important part of the antibacterial armamentarium. Here, in order to identify novel inhibitors of bacterial RNAP, we used affinity-selection mass spectrometry to screen a chemical library for compounds that bind to Escherichia coli RNAP. We identified a novel small molecule, MRL-436, that binds to RNAP, inhibits RNAP, and exhibits antibacterial activity. MRL-436 binds to RNAP through a binding site that differs from the rifampin binding site, inhibits rifampin-resistant RNAP derivatives, and exhibits antibacterial activity against rifampin-resistant strains. Isolation of mutants resistant to the antibacterial activity of MRL-436 yields a missense mutation in codon 622 of the rpoC gene encoding the RNAP β' subunit or a null mutation in the rpoZ gene encoding the RNAP ω subunit, confirming that RNAP is the functional cellular target for the antibacterial activity of MRL-436, and indicating that RNAP β' subunit residue 622 and the RNAP ω subunit are required for the antibacterial activity of MRL-436. Similarity between the resistance determinant for MRL-436 and the resistance determinant for the cellular alarmone ppGpp suggests a possible similarity in binding site and/or induced conformational state for MRL-436 and ppGpp.
Merck & Co., Inc. , Kenilworth, New Jersey 07033, United States.
Scott S
0000-0002-3076-439X
SS
Walker
Department of Chemistry and Waksman Institute, Rutgers University , Piscataway, New Jersey 08854, United States.
David
D
Degen
Merck & Co., Inc. , Boston, Massachusetts 02115, United States.
Elliott
E
Nickbarg
Merck & Co., Inc. , Kenilworth, New Jersey 07033, United States.
Donna
D
Carr
Merck & Co., Inc. , Kenilworth, New Jersey 07033, United States.
Aileen
A
Soriano
Merck & Co., Inc. , Kenilworth, New Jersey 07033, United States.
Mihir
0000-0002-7870-7271
M
Mandal
Merck & Co., Inc. , Kenilworth, New Jersey 07033, United States.
Ronald E
RE
Painter
Merck & Co., Inc. , Kenilworth, New Jersey 07033, United States.
Payal
P
Sheth
Merck & Co., Inc. , Kenilworth, New Jersey 07033, United States.
Li
L
Xiao
Merck & Co., Inc. , Boston, Massachusetts 02115, United States.
Xinwei
X
Sher
Merck & Co., Inc. , Kenilworth, New Jersey 07033, United States.
Nicholas
N
Murgolo
Merck & Co., Inc. , Kenilworth, New Jersey 07033, United States.
Jing
J
Su
Merck & Co., Inc. , Upper Gwynedd, Pennsylvania 19454, United States.
David B
DB
Olsen
Department of Chemistry and Waksman Institute, Rutgers University , Piscataway, New Jersey 08854, United States.
Richard H
RH
Ebright
Merck & Co., Inc. , Kenilworth, New Jersey 07033, United States.
Katherine
K
Young
D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001665:Binding Sites; D012321:DNA-Directed RNA Polymerases / Q000037:antagonists & inhibitors*; D024881:Drug Resistance, Bacterial / Q000187:drug effects* / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology; D013058:Mass Spectrometry; D011485:Protein Binding; D012293:Rifampin / Q000494:pharmacology; D054852:Small Molecule Libraries
D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D054852:Small Molecule Libraries; D012321:DNA-Directed RNA Polymerases; D012293:Rifampin