Do acyl homoserine lactones (AHLs) compete with endogenous immunomodulators by acting as substrates for cyp450?

Abstract

Pseudomonas aeruginosa (PA) is a major cause of fatalities in Cystic Fibrosis (CF) patients due to the virulence factors such as pyocyanin and toxins like cyanide that this organism produces. The expression of virulent genes occurs when the lipid signal acyl homoserine lactone (AHL) binds to transcription factors expressing a particular phenotype. Quorum sensing (QS) is the term given to the ability of bacteria to sense their own population density and therefore express certain phenotypes. QS is activated when AHLs reach a threshold concentration. The QSS‐regulated genes play an important role in the persistence of PA during the early infection phase. Preliminary data has shown that P450BM‐3 interacts with and can initiate inactivation of some AHLs. P450BM‐3 shares sequence homology with human P450s, including P450s that regulate immunomodulators such as leukotriene B4 (LTB4). LTB4 plays an important role in neutrophil trafficking and has been identified in large numbers in lung tissue of CF patients, and if the P450s that metabolize it also metabolize AHLs there is the potential for AHL‐mediated interference with immune regulation via this interaction. We are identifying the metabolites and which specific P450s in HLM are responsible for the hydroxylation of the AHLs. Assays were performed using human liver microsomes, NADPH and AHL but there is a need for hydroxylated AHL standards to confirm our results. Hydroxylated AHL standards where produced through an enzymatic reaction using a promiscuous mutant of P450BM‐3 in an aqueous turnover system. AHLs and their oxidation products were analyzed by GC/MS. Want to give thanks to the Robert A. Welch Foundation for the support.