The structure and function of esterases from lactic acid bacteria : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosphy in the Institute of Molecular BioSciences, Massey University, New Zealand
| dc.contributor.author | Bennett, Matthew David | |
| dc.date.accessioned | 2009-05-06T21:34:10Z | |
| dc.date.available | NO_RESTRICTION | en_US |
| dc.date.available | 2009-05-06T21:34:10Z | |
| dc.date.issued | 2007 | |
| dc.description.abstract | Compounds derived from the breakdown of glyceride esters of milk fat, such as free fatty acids and short chain esters, are recognised as playing an important role in the flavour of a range of fermented foods. Esterases, capable of hydrolysing ester bonds, and in some cases, synthesising them via an acyltransferase mechanism, typically enter the fermentation from the starter and adjunct lactic acid bacteria that are used to inoculate milk to initiate the fermentation process. With such an important role in the development of both desirable and undesirable flavours, understanding how these enzymes operate is essential for product control. In this study, the crystal structures of three lactic acid bacterial esterases were solved: EstA from Lactococcus lactis, and AA7 from Lactobacillus rhamnosus which are both capable of hydrolysis of short chain triglycerides as well as synthesising esters via a transferase mechanism, and AZ4, an esterase from L. rhamnosus which appears to be limited to hydrolysis reactions. Whilst all three were found to be members of the hydrolase family, unique features were found for each enzyme, reflecting the large differences in their primary sequences, substrate specificities and activities. EstA and AA7 were both found to have a shallow substrate binding cleft, bisected by the catalytic machinery. The divided binding cleft suggests that during a transferase reaction the transferred group binds in one pocket, with the donor and acceptor groups (dependant on the stage of catalysis) binding in the other. In contrast, AZ4 was found to have a single deep substrate binding cavity, extending into the enzyme interior, with the catalytic residues located near its entrance. The absence of a second binding site for an acceptor is consistent with AZ4 having only one function – that of a hydrolase. The structures presented in this study are the first three dimensional structures of esterases from lactic acid bacteria to be reported. Their analyses, both in native form, and complexed with a varity of ligands mimicking various stages of the reaction cycle have highlighted how this basic fold can be adapted to efficiently catalyse different reactions. More importantly, in the case of AZ4, these structures have suggested that there is a novel mechanism used by the esterases to promote the enzyme reaction to proceed to completion, by preventing a futile catalytic reaction. | en_US |
| dc.identifier.uri | http://hdl.handle.net/10179/810 | |
| dc.language.iso | en | en_US |
| dc.publisher | Massey University | en_US |
| dc.rights | The Author | en_US |
| dc.subject | Lactic acid | en_US |
| dc.subject | Milk | en_US |
| dc.subject | Esterases | en_US |
| dc.subject.other | Fields of Research::270000 Biological Sciences::270100 Biochemistry and Cell Biology::270109 Structural biology | en_US |
| dc.title | The structure and function of esterases from lactic acid bacteria : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosphy in the Institute of Molecular BioSciences, Massey University, New Zealand | en_US |
| dc.type | Thesis | en_US |
| massey.contributor.author | Bennett, Matthew David | |
| thesis.degree.discipline | Biochemistry | en_US |
| thesis.degree.grantor | Massey University | en_US |
| thesis.degree.level | Doctoral | en_US |
| thesis.degree.level | Doctoral | en |
| thesis.degree.name | Doctor of Philosophy (Ph.D.) | en_US |
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