Quorum quenching effect of recombinant Paraoxonase-1 enzyme against quorum sensing genes produced from Pseudomonas aeruginosa

Abstract

Inhibition of quorum sensing (QS), also known as quorum quenching (QQ), is thought to provide means of fighting Pseudomonas aeruginosa infections, Paraoxonase-1 (PON1) enzymes has been selected as a candidate for inhibiting QS signaling activity of P. aeruginosa. This study was conducted to produce a recombinant human PON1 (hPON1) in a bacterial host and evaluate its QQ affect against QS genes produced from P. aeruginosa. Circulating lactonase activity of PON1 were measured in the 50 serum samples, the PON1 gene was amplified by using a NOVA Taq DNA polymerase. After RT-PCR, the purified restricted PCR product hPON1 gene was ligated into pETBlue-1 by utilizing Clonables™ 2× Ligation Premix. Transformation to Nova Blue Singles™ Competent Cells and Tuner (DE3) pLacI was done by using (heat shock) method and the expression of QS gene was measured by using KAPA SYBR FAST qPCR. Results revealed that hPON1 gene was successfully cloned in to pETBlue-1 and white colonies of Nova Blue E. coli recombinant and was observed in Luria-Bertani (LB) agar plates contain IPTG as an inducer and X-gal as an indicator, the relative gene expression data obtained showed loss of AHL concentration by about 14-fold reduction in lasI gene expression while lasR showed 3-fold reduction when compared to the untreated cells. The reduction in rhlI gene synthase following recombinant PON1 treatment by 6-fold while in rhIR gene synthase was 3- fold reduction when compared to the untreated cells. Our study showed that, in addition of being generally effective, hPON1 could be a suitable candidate as a QQ agent sufficient to inhibit the virulence factors produced by P. aeruginosa.