Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.716422
Title: The metabolism of plant glucosinolates by gut bacteria
Author: Cebeci, Fatma
Awarding Body: University of East Anglia
Current Institution: University of East Anglia
Date of Award: 2017
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Abstract:
Glucosinolates found in cruciferous vegetables are degraded by plant myrosinases into bioactive isothiocyanates (ITCs) which have been recognised as potent anticancer compounds. During cooking, plant myrosinases are heat inactivated so ITC production is dependent on the myrosinase-like enzymes produced by the gut bacteria. This study is focused on investigating glucosinolate metabolism by the human gut bacteria and identifying the enzymes that play a crucial role. Human gut bacteria that were previously reported to metabolise glucosinolates were investigated in this study. In addition, 98 more human gut strains were isolated using a glucoraphanin enrichment method. It was hypothesised that bacterial myrosinases are β-glucosidases with specificity for glucosinolates. To identify the first bacterial myrosinase from the human gut, four putative β-glucosidases from Enterococcus casseliflavus CP1 and Escherichia coli FI10944 were cloned and heterologously expressed in E. coli. An alternative approach using a combination of ion exchange chromatography and gel filtration was also carried out to identify the bacterial myrosinase of E. coli FI109444. It has been reported that some gut bacteria require a reduction step to metabolise methylsulfinylalkyl glucosinolates (such as glucoraphanin) that converts them into methylthioalkyl glucosinolates (such as glucoerucin) to produce ITCs. To identify the responsible reductase, candidate reductase genes were cloned and expressed in E. coli. Methionine sulfoxide reductase B (MsrB) from Escherichia coli VL8 and Lactobacillus agilis R16 was found to reduce glucoraphanin to glucoerucin under the conditions tested. A bacterial myrosinase of Citrobacter WYE1 of soil origin was previously identified and myrosinase activity of this enzyme was characterised using cell-free extracts. In this study the myrosinase gene was heterologously expressed in E. coli to allow purification and characterisation. The recombinant enzyme showed activity against several glucosinolate substrates and protein was produced for crystallographic studies.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.716422  DOI: Not available
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