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Allosteric Competitive Inhibitors of the Glucose-1-phosphate Thymidylyltransferase (RmlA) from Pseudomonas aeruginosa

DOI: 10.1021/cb300426u DOI Help
PMID: 23138692 PMID Help

Authors: Magnus Alphey (University of St. Andrews) , Lisa Pirrie (University of St. Andrews) , Leah S. Torrie (University of Dundee) , Wassila Abdelli Boulkeroua (University of St. Andrews) , Mary Gardiner (University of Dundee) , Aurijit Sarkar (University of Dundee) , Marko Maringer (mfd Diagnostics GmbH) , Wulf Oehlmann (Lionex GmbH) , Ruth Brenk (University of Dundee) , Michael S. Scherman (Colorado State University) , Michael Mcneil (Colorado State University) , Martin Rejzek (John Innes Centre) , Robert A. Field (John Innes Centre) , Mahavir Singh (Lionex GmbH) , David Gray (University of Dundee) , Nicholas J. Westwood (University of St. Andrews) , James H. Naismith (University of St. Andrews)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Acs Chemical Biology , VOL 8 (2)

State: Published (Approved)
Published: November 2012

Open Access Open Access

Abstract: Glucose-1-phosphate thymidylyltransferase (RmlA) catalyzes the condensation of glucose-1-phosphate (G1P) with deoxy-thymidine triphosphate (dTTP) to yield dTDP-d-glucose and pyrophosphate. This is the first step in the l-rhamnose biosynthetic pathway. l-Rhamnose is an important component of the cell wall of many microorganisms, including Mycobacterium tuberculosis and Pseudomonas aeruginosa. Here we describe the first nanomolar inhibitors of P. aeruginosa RmlA. These thymine analogues were identified by high-throughput screening and subsequently optimized by a combination of protein crystallography, in silico screening, and synthetic chemistry. Some of the inhibitors show inhibitory activity against M. tuberculosis. The inhibitors do not bind at the active site of RmlA but bind at a second site remote from the active site. Despite this, the compounds act as competitive inhibitors of G1P but with high cooperativity. This novel behavior was probed by structural analysis, which suggests that the inhibitors work by preventing RmlA from undergoing the conformational change key to its ordered bi-bi mechanism.

Journal Keywords: Binding; Competitive; Enzyme; High-Throughput; Models; Molecular; Nucleotidyltransferases; Pseudomonas; Structure-Activity; Thymine

Subject Areas: Biology and Bio-materials, Chemistry


Instruments: I02-Macromolecular Crystallography , I03-Macromolecular Crystallography , I04-1-Macromolecular Crystallography (fixed wavelength) , I04-Macromolecular Crystallography , I24-Microfocus Macromolecular Crystallography