Aii810, a Novel Cold-Adapted N-Acylhomoserine Lactonase Discovered in a Metagenome, Can Strongly Attenuate Pseudomonas aeruginosa Virulence Factors and Biofilm Formation.

Xinjiong Fan,He Li,Ruo Chen,Mingjun Liang,Xiaolong Liu,Lei Wang

doi:10.3389/fmicb.2017.01950

Xinjiong Fan, He Li + Show 4 more

Open Access

https://doi.org/10.3389/fmicb.2017.01950

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Abstract

The pathogen Pseudomonas aeruginosa uses quorum sensing (QS) to control virulence and biofilm formation. Enzymatic disruption of quorum sensing is a promising anti-infection therapeutic strategy that does not rely on antibiotics. Here, a novel gene (aii810) encoding an N-acylhomoserine lactonase was isolated from the Mao-tofu metagenome for the first time. Aii810 encoded a protein of 269 amino acids and was expressed in Escherichia coli BL21 (DE3) in soluble form. It showed the highest activity at 20°C, and it maintained 76.5% of activity at 0°C and more than 50% activity at 0–40°C. The optimal pH was 8.0. It was stable in both neutral and slightly alkaline conditions and at temperatures below 40°C. The enzyme hydrolyzed several ρ-nitrophenyl esters, but its best substrate was ρ-nitrophenyl acetate. Its kcat and Km values were 347.7 S-1 and 205.1 μM, respectively. It efficiently degraded N-butyryl-L-homoserine lactone and N-(3-oxododecanoyl)-L-homoserine lactone, exceeding hydrolysis rates of 72.3 and 100%, respectively. Moreover, Aii810 strongly attenuated P. aeruginosa virulence and biofilm formation. This enzyme with high anti-QS activity was the most cold-adapted N-acylhomoserine lactonase reported, which makes it an attractive enzyme for use as a therapeutic agent against P. aeruginosa infection.

Highlights

Pseudomonas aeruginosa is a Gram-negative bacterium that thrives in diverse terrestrial and aquatic environments
The inhibition of biofilm formation against P. aeruginosa PAO1 was characterized by scanning electron microscopy (SEM) as described by Grover et al (2016)
These results suggest that Aii810 has high activity toward these two acyl L-homoserine lactones (AHLs) and possesses good adaptability at moderate and low temperatures, which may be useful in targeting the quorum sensing (QS) systems in P. aeruginosa, but these findings needs further exploration

Summary

INTRODUCTION

Pseudomonas aeruginosa is a Gram-negative bacterium that thrives in diverse terrestrial and aquatic environments. The procedure relies on the production, release, and group-wide detection of small diffusible signal molecules, which are typically N-acyl L-homoserine lactones (AHLs) in gram-negative bacteria. P. aeruginosa possesses two QS systems based on acyl homoserine lactone molecules to orchestrate synchronous production of biofilm formation and virulence factors (de Kievit, 2009). We reported the molecular cloning, identification, and biochemical characterization of one novel cold-adapted N-acylhomoserine lactonase derived from the Mao-tofu metagenome. This enzyme showed high activity at moderate and low temperatures. It efficiently degraded BHL and OdDHL, and significantly decreased the production of virulence factors and biofilm in P. aeruginosa PAO1

MATERIALS AND METHODS

RESULTS AND DISCUSSION

CONCLUSION