title: Rational design of a transition state analogue with picomolar affinity for Pseudomonas aeruginosa PvdQ, a siderophore biosynthetic enzyme.
abstract: The Pseudomonas aeruginosa enzyme PvdQ can process different substrates involved in quorum-sensing or in siderophore biosynthesis. Substrate selectivity was evaluated using steady-state kinetic constants for hydrolysis of N-acyl-homoserine lactones (HSLs) and p-nitrophenyl fatty acid esters. PvdQ prefers substrates with alkyl chains between 12 and 14 carbons long that do not bear a 3-oxo substitution and is revealed here to have a relatively high specificity constant for selected N-acyl-HSLs (kcat/KM = 10(5) to 10(6) M(-1) s(-1)). However, endogenous P. aeruginosa N-acyl-HSLs are ≥100-fold disfavored, supporting the conclusion that PvdQ was not primarily evolved to regulate endogenous quorum-sensing. PvdQ plays an essential biosynthetic role for the siderophore pyoverdine, on which P. aeruginosa depends for growth in iron-limited environments. A series of alkylboronate inhibitors was found to be reversible, competitive, and extremely potent (Ki ≥ 190 pM). A 1.8 Å X-ray structure shows that 1-tridecylboronic acid forms a monocovalent bond with the N-terminal β-chain Ser residue in the PvdQ heterodimer, mimicking a reaction transition state. This boronic acid inhibits growth of P. aeruginosa in iron-limited media, reproducing the phenotype of a genetic pvdQ disruption, although co-administration of an efflux pump inhibitor is required to maintain growth inhibition. These findings support the strategy of designing boron-based inhibitors of siderophore biosynthetic enzymes to control P. aeruginosa infections.
authors.affiliation: Department of Chemistry and Biochemistry and the ‡College of Pharmacy, Medicinal Chemistry Division, University of Texas , Austin, Texas 78712, United States.
authors.forename: Kenneth D
authors.initials: KD
authors.lastname: Clevenger
authors.forename: Rui
authors.initials: R
authors.lastname: Wu
authors.forename: Joyce A V
authors.initials: JA
authors.lastname: Er
authors.forename: Dali
authors.initials: D
authors.lastname: Liu
authors.forename: Walter
authors.initials: W
authors.lastname: Fast
mesh_terms: D001681:Biological Assay; D002626:Chemistry, Pharmaceutical; D018360:Crystallography, X-Ray; D015195:Drug Design; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology; D020128:Inhibitory Concentration 50; D008024:Ligands; D008958:Models, Molecular; D011550:Pseudomonas aeruginosa / Q000187:drug effects / Q000201:enzymology*; D017262:Siderophores / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D004791:Enzyme Inhibitors; D008024:Ligands; D017262:Siderophores