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Investigation of a general base mechanism for ester hydrolysis in C-C hydrolase enzymes of the alpha/beta-hydrolase superfamily : a novel mechanism for the serine catalytic triad
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Li, Jian-Jun and Bugg, Tim (2007) Investigation of a general base mechanism for ester hydrolysis in C-C hydrolase enzymes of the alpha/beta-hydrolase superfamily : a novel mechanism for the serine catalytic triad. Organic & Biomolecular Chemistry, Volume 5 (Number 3). pp. 507-513. doi:10.1039/b615605c
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Official URL: http://dx.doi.org/10.1039/b615605c
Abstract
Previous mechanistic and crystallographic studies on two C - C hydrolase enzymes, Escherichia coli MhpC and Burkholderia xenovorans BphD, support a general base mechanism for C - C hydrolytic cleavage, rather than the nucleophilic mechanism expected for a serine hydrolase. The role of the active site serine residue could be to form a hydrogen bond with a gem-diolate intermediate, or to protonate such an intermediate. Hydrolase BphD is able to catalyse the hydrolysis of p-nitrophenyl benzoate ester substrates, which has enabled an investigation of these mechanisms using a Hammett analysis, and comparative studies upon five serine esterases and lipases from the alpha/beta-hydrolase family. A reaction parameter (rho) value of + 0.98 was measured for BphD-catalysed ester hydrolysis, implying a build-up of negative charge in the transition state, consistent with a general base mechanism. Values of + 0.31 - 0.61 were measured for other serine esterases and lipases, for the same series of esterase substrates. Pre-steady state kinetic studies of ester hydrolysis, using p-nitrophenyl acetate as the substrate, revealed a single step kinetic mechanism for BphD-catalysed ester hydrolysis, with no burst kinetics. A general base mechanism for BphD-catalysed ester hydrolysis is proposed, in which Ser-112 stabilises an oxyanion intermediate through hydrogen bonding, and assists the rotation of this oxyanion intermediate via proton transfer, a novel reaction mechanism for the serine catalytic triad.
| Item Type: | Journal Article | ||||
|---|---|---|---|---|---|
| Subjects: | Q Science > QD Chemistry | ||||
| Divisions: | Faculty of Science > Chemistry | ||||
| Journal or Publication Title: | Organic & Biomolecular Chemistry | ||||
| Publisher: | Royal Society of Chemistry | ||||
| ISSN: | 1477-0520 | ||||
| Official Date: | 2007 | ||||
| Dates: |
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| Volume: | Volume 5 | ||||
| Number: | Number 3 | ||||
| Number of Pages: | 7 | ||||
| Page Range: | pp. 507-513 | ||||
| DOI: | 10.1039/b615605c | ||||
| Status: | Peer Reviewed | ||||
| Publication Status: | Published |
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