Structure of an aryl esterase from<i>Pseudomonas fluorescens</i>

Cheeseman, Jeremy D.; Tocilj, Ante; Park, Seongsoon; Schrag, Joseph D.; Kazlauskas, Romas J.

doi:10.1107/s0907444904010522

Cited by 70 publications

(80 citation statements)

References 23 publications

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“…Similarly, the alcohol pocket in the PfEst structure is lined with several hydrophobic phenylalanine side chains that should have affinity for the lactone ring. This would explain its lactonase activity towards caprolactone [27] in addition to the esterase activity.…”

Section: Structural Comparisonmentioning

confidence: 99%

Structural studies of a thermophilic esterase from a new Planctomycetes species, Thermogutta terrifontis

et al. 2015

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Thermogutta terrifontis esterase (TtEst), a carboxyl esterase identified in the novel thermophilic bacterium T. terrifontis from the phylum Planctomycetes, has been cloned and over-expressed in Escherichia coli. The enzyme has been characterized biochemically and shown to have activity towards small p-nitrophenyl (pNP) carboxylic esters, with optimal activity for pNPpropionate. The enzyme retained 95% activity after incubation for 1 h at 80°C. The crystal structures of the native TtEst and its complexes with the substrate analogue D-malate and the product acetate have been determined to high resolution. The bound ligands have allowed the identification of the carboxyl and alcohol binding pockets in the enzyme active site. Comparison of TtEst with structurally related enzymes provides insight into how differences in their catalytic activity can be rationalized based upon the properties of the amino acid residues in their active site pockets. The mutant enzymes L37A and L251A have been constructed to extend the substrate range of TtEst towards the larger butyrate and valerate pNPesters. These mutant enzymes have also shown a significant increase in activity towards acetate and propionate pNP esters. A crystal structure of the L37A mutant was determined with the butyrate product bound in the carboxyl pocket of the active site. The mutant structure shows an expansion of the pocket that binds the substrate carboxyl group, which is consistent with the observed increase in activity towards pNP-butyrate.

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Section: Structural Comparisonmentioning

confidence: 99%

Structural studies of a thermophilic esterase from a new Planctomycetes species, Thermogutta terrifontis

et al. 2015

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“…6 In situ generation of peroxycarboxylic acids by hydrolysis of esters or amides in the presence of hydrogen peroxide is an alternative approach for producing active peroxycarboxylic acids when required in applications. 7 Peroxycarboxylic acid production employing hydrolases has been reported for proteases, 8,9 lipases, 10 and esterases, 11 which are mainly used in detergent formulations. In a patent application, 8 the reengineering of proteases for peroxycarboxylic acid production was reported for the first time.…”

Section: Introductionmentioning

confidence: 99%

Fluorescent Assay for Directed Evolution of Perhydrolases

et al. 2012

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Directed evolution offers opportunities to improve promiscuous activities of hydrolases in rounds of diversity generation and high-throughput screening. In this article, we developed and validated a screening platform to improve the perhydrolytic activity of proteases and likely other hydrolases (e.g., lipases or esterases). Key was the development of a highly sensitive fluorescent assay (sensitivity in the µM range) based on 3-carboxy-7-hydroxycoumarin (HCC) formation. HCC is released through an hypobromite-mediated oxidation of 7-(4′-aminophenoxy)-3-carboxycoumarin (APCC), which enables for the first time a continuous measurement of peroxycarboxylic acid formation with a standard deviation of 11% in microtiter plates with a wide pH range window (5-9). As example, subtilisin Carlsberg was subjected to site saturation mutagenesis at position G165, yielding a variant T58A/G165L/L216W with 5.4-fold increased k cat for perhydrolytic activity compared with wild type.

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“…Interestingly, the distribution of secondary structures in AAMS amidohydrolase and aryl esterase is quite similar, suggesting that they have a similar tertiary structure. The aryl esterase, which is a metal-independent hydrolase, has the catalytic triad (Ser94, Asp222, and His251) (17) which is shown in red colors in Fig. 3.…”

Section: Amino Acid Sequence Comparisonsmentioning

confidence: 99%

“…Then, the sequence was analyzed by BLAST: the enzyme showed no identity to amidohydrolases so far reported. However, it showed low but significant (about 27%) homology to fluoroacetate dehalogenase (PDB code, 1Y37) (15), haloalkane dehalogenase (1K5P) (16), and aryl esterase (1VA4) (17). Amino acid sequences of eleven hydrolases, whose substrate specificity is characterized, were aligned by CLUSTALW, and an unrooted tree was determined (Fig.…”

Section: Amino Acid Sequence Comparisonsmentioning

confidence: 99%

Gene Identification and Characterization of the Pyridoxine Degradative Enzyme .ALPHA.-(N-Acetylaminomethylene)succinic Acid Amidohydrolase from Mesorhizobium loti MAFF303099

Yuan

Yokochi

Yoshikane

et al. 2008

J Nutr Sci Vitaminol

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SummaryWe have found for the first time that a chromosomal gene, mlr6787, in Mesorhizobium loti encodes the pyridoxine degradative enzyme ␣ -( N -acetylaminomethylene)succinic acid (AAMS) amidohydrolase. The recombinant enzyme expressed in Escherichia coli cells was homogeneously purified and characterized. The enzyme consisted of two subunits each with a molecular mass of 34,000 Ϯ 1,000 Da, and exhibited Km and kcat values of 53.7 Ϯ 6 M and 307.3 Ϯ 12 min Ϫ 1 , respectively. The enzyme required no cofactor or metal ion. The primary structure of AAMS amidohydrolase was elucidated for the first time here. The primary structure of the enzyme protein showed no significant identity to those of known hydrolase proteins and low homology to those of fluoroacetate dehalogenase (PDB code, 1Y37), haloalkane dehalogenase (1K5P), and aryl esterase (1VA4).

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Structure of an aryl esterase fromPseudomonas fluorescens

Cited by 70 publications

References 23 publications

Structural studies of a thermophilic esterase from a new Planctomycetes species, Thermogutta terrifontis

Structural studies of a thermophilic esterase from a new Planctomycetes species, Thermogutta terrifontis

Fluorescent Assay for Directed Evolution of Perhydrolases

Gene Identification and Characterization of the Pyridoxine Degradative Enzyme .ALPHA.-(N-Acetylaminomethylene)succinic Acid Amidohydrolase from Mesorhizobium loti MAFF303099

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