Abstract
Many Gram-negative bacteria produce N-acyl-homoserine lactones (AHLs), quorum sensing (QS) molecules that can be enzymatically inactivated by quorum quenching (QQ) processes; this approach is considered an emerging antimicrobial alternative. In this study, kinetic parameters of several AHLs hydrolyzed by penicillin acylase from Streptomyces lavendulae (SlPA) and aculeacin A acylase from Actinoplanes utahensis (AuAAC) have been determined. Both enzymes catalyze efficiently the amide bond hydrolysis in AHLs with different acyl chain moieties (with or without 3-oxo modification) and exhibit a clear preference for AHLs with long acyl chains (C12-HSL > C14-HSL > C10-HSL > C8-HSL for SlPA, whereas C14-HSL > C12-HSL > C10-HSL > C8-HSL for AuAAC). Involvement of SlPA and AuAAC in QQ processes was demonstrated by Chromobacterium violaceum CV026-based bioassays and inhibition of biofilm formation by Pseudomonas aeruginosa, a process controlled by QS molecules, suggesting the application of these multifunctional enzymes as quorum quenching agents, this being the first time that quorum quenching activity was shown by an aculeacin A acylase. In addition, a phylogenetic study suggests that SlPA and AuAAC could be part of a new family of actinomycete acylases, with a preference for substrates with long aliphatic acyl chains, and likely involved in QQ processes.
Highlights
Quorum sensing (QS) is a bacterial cell-to-cell communication mechanism that allows bacteria to regulate a high diversity of biological functions such as bioluminescence, production of virulence factors, antibiotics and other secondary metabolites, and biofilm formation by releasing, detecting and responding to small, diffusible signal molecules called autoinducers [1,2]
Our study demonstrates that two other enzymes from Gram-positive bacteria, such as SlPA and AuAAC, present this feature, showing penicillin acylase and acyl-homoserine lactones (AHLs) acylase activities, and aculeacin
We have demonstrated that reported enzymes, penicillin acylase from Streptomyces lavendulae and aculeacin A acylase from Actinoplanes utahensis, are an interesting extension of hydrolytic (Ntn) enzymes, with potential for biocatalytic applications
Summary
Quorum sensing (QS) is a bacterial cell-to-cell communication mechanism that allows bacteria to regulate a high diversity of biological functions such as bioluminescence, production of virulence factors, antibiotics and other secondary metabolites, and biofilm formation by releasing, detecting and responding to small, diffusible signal molecules called autoinducers [1,2]. The Ntn-hydrolase superfamily contains many enzymes with diverse activities, including β-lactam acylases, AHL acylases and proteasomes, among others [17] Many of these enzymes have been classified according to their respective first reported activities, this classification is not always necessarily in agreement with their true biological role [18]. (AuAAC) (EC 3.5.1.70) are the unique described penicillin and echinocandin acylases, respectively, capable of efficiently hydrolyzing phenoxymethyl penicillin (penicillin V), several natural aliphatic penicillins (such as penicillin K, penicillin F and penicillin dihydro F) and aculeacin A [21,22,23,24] Both show very similar substrate specificity, with a marked preference for amides bearing long hydrophobic acyl moieties [21,22,24]. High identities between both enzymes and AHL quorum quenching acylases were revealed by comparative sequence analysis
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