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Biochemical studies on a versatile esterase that is most catalytically active with polyaromatic esters.

ABSTRACT: Herein, we applied a community genomic approach using a naphthalene-enriched community (CN1) to isolate a versatile esterase (CN1E1) from the ?/?-hydrolase family. The protein shares low-to-medium identity (??57%) with known esterase/lipase-like proteins. The enzyme is most active at 25-30°C and pH?8.5; it retains approximately 55% of its activity at 4°C and less than 8% at ??55°C, which indicates that it is a cold-adapted enzyme. CN1E1 has a distinct substrate preference compared with other ?/?-hydrolases because it is catalytically most active for hydrolysing polyaromatic hydrocarbon (phenanthrene, anthracene, naphthalene, benzoyl, protocatechuate and phthalate) esters (7200-21?000?units?g(-1) protein at 40°C and pH?8.0). The enzyme also accepts 44 structurally different common esters with different levels of enantio-selectivity (1.0-55?000?units?g(-1) protein), including (±)-menthyl-acetate, (±)-neomenthyl acetate, (±)-pantolactone, (±)-methyl-mandelate, (±)-methyl-lactate and (±)-glycidyl 4-nitrobenzoate (in that order). The results provide the first biochemical evidence suggesting that such broad-spectrum esterases may be an ecological advantage for bacteria that mineralize recalcitrant pollutants (including oil refinery products, plasticizers and pesticides) as carbon sources under pollution pressure. They also offer a new tool for the stereo-assembly (i.e. through ester bonds) of multi-aromatic molecules with benzene rings that are useful for biology, chemistry and materials sciences for cases in which enzyme methods are not yet available.

SUBMITTER: Martinez-Martinez M

PROVIDER: S-EPMC3937722 | biostudies-literature | 2014 Mar

REPOSITORIES: biostudies-literature

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Biochemical studies on a versatile esterase that is most catalytically active with polyaromatic esters.

Martínez-Martínez Mónica M Lores Iván I Peña-García Carlina C Bargiela Rafael R Reyes-Duarte Dolores D Guazzaroni María-Eugenia ME Peláez Ana Isabel AI Sánchez Jesús J Ferrer Manuel M

Microbial biotechnology 20140113 2

Herein, we applied a community genomic approach using a naphthalene-enriched community (CN1) to isolate a versatile esterase (CN1E1) from the α/β-hydrolase family. The protein shares low-to-medium identity (≤ 57%) with known esterase/lipase-like proteins. The enzyme is most active at 25-30°C and pH 8.5; it retains approximately 55% of its activity at 4°C and less than 8% at ≥ 55°C, which indicates that it is a cold-adapted enzyme. CN1E1 has a distinct substrate preference compared with other α/β ...[more]

PMID: 24418210

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Project description:Pseudomonas aeruginosa is a human pathogen that relies on quorum sensing to establish infections. The PqsE quorum-sensing protein is required for P. aeruginosa virulence factor production and infection. PqsE has a reported enzymatic function in the biosynthesis of the quorum-sensing autoinducer called PQS. However, this activity is redundant because, in the absence of PqsE, this role is fulfilled by alternative thioesterases. Rather, PqsE drives P. aeruginosa pathogenic traits via a protein-protein interaction with the quorum-sensing receptor/transcription factor RhlR, an interaction that enhances the affinity of RhlR for target DNA sequences. PqsE catalytic activity is dispensable for interaction with RhlR. Thus, the virulence function of PqsE can be decoupled from its catalytic function. Here, we present an immunoprecipitation-mass spectrometry method employing enhanced green fluorescent protein-PqsE fusions to define the protein interactomes of wild-type PqsE and the catalytically inactive PqsE(D73A) variant in P. aeruginosa and their dependence on RhlR. Several proteins were identified to have specific interactions with wild-type PqsE while not forming associations with PqsE(D73A). In the ΔrhlR strain, an increased number of specific PqsE interactors were identified, including the partner autoinducer synthase for RhlR, called RhlI. Collectively, these results suggest that specific protein-protein interactions depend on PqsE catalytic activity and that RhlR may prevent proteins from interacting with PqsE, possibly due to competition between RhlR and other proteins for PqsE binding. Our results provide a foundation for the identification of the in vivo PqsE catalytic function and, potentially, new proteins involved in P. aeruginosa quorum sensing. IMPORTANCE Pseudomonas aeruginosa causes hospital-borne infections in vulnerable patients, including immunocompromised individuals, burn victims, and cancer patients undergoing chemotherapy. There are no effective treatments for P. aeruginosa infections, which are usually broadly resistant to antibiotics. Animal models show that, to establish infection and to cause illness, P. aeruginosa relies on an interaction between two proteins, namely, PqsE and RhlR. There could be additional protein-protein interactions involving PqsE, which, if defined, could be exploited for the design of new therapeutic strategies to combat P. aeruginosa. Here, we reveal previously unknown protein interactions in which PqsE participates, which will be investigated for potential roles in pathogenesis.

Dataset Information

Biochemical studies on a versatile esterase that is most catalytically active with polyaromatic esters.

Publications

Biochemical studies on a versatile esterase that is most catalytically active with polyaromatic esters.

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