Structural and Functional Analysis of Grapefruit Flavonol-Specific-3-O-GT Mutant P145T
This research is focused on the study of the effect of mutating proline 145 to threonine on the substrate and regiospecificity of flavonol specific 3-O-glucosyltransferase (Cp3GT). While the mutant P145T enzyme did not glucosylate anthocyanidins, it did glucosylate flavanones and flavones in additio...
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ndltd-ETSU-oai-dc.etsu.edu-etd-45602019-05-16T05:07:16Z Structural and Functional Analysis of Grapefruit Flavonol-Specific-3-O-GT Mutant P145T Kandel, Sangam, Mr This research is focused on the study of the effect of mutating proline 145 to threonine on the substrate and regiospecificity of flavonol specific 3-O-glucosyltransferase (Cp3GT). While the mutant P145T enzyme did not glucosylate anthocyanidins, it did glucosylate flavanones and flavones in addition to retaining activity with flavonols. HPLC was used for product identification and showed mutant P145T glucosylated naringenin at the 7-OH position forming naringenin-7-O-glucoside and flavonols at the 3-OH position. Homology modeling and docking was done to predict the acceptor substrate recognition pattern and models were validated by experimental results. In other related work, a thrombin cleavage site was inserted into wild type Cp3GT and recombinant P145T enzyme between the enzyme and the C-myc tags in order to be able to cleave off tags. This provides the tool needed for future efforts to crystallize these proteins for structural determination. 2016-12-01T08:00:00Z text application/pdf https://dc.etsu.edu/etd/3150 https://dc.etsu.edu/cgi/viewcontent.cgi?article=4560&context=etd Copyright by the authors. Electronic Theses and Dissertations eng Digital Commons @ East Tennessee State University Glucosyltrasnferase Citrus paradisi Mutation Flavonoids Glucosides Homology modeling Biology |
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English |
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Others
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Glucosyltrasnferase Citrus paradisi Mutation Flavonoids Glucosides Homology modeling Biology |
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Glucosyltrasnferase Citrus paradisi Mutation Flavonoids Glucosides Homology modeling Biology Kandel, Sangam, Mr Structural and Functional Analysis of Grapefruit Flavonol-Specific-3-O-GT Mutant P145T |
description |
This research is focused on the study of the effect of mutating proline 145 to threonine on the substrate and regiospecificity of flavonol specific 3-O-glucosyltransferase (Cp3GT). While the mutant P145T enzyme did not glucosylate anthocyanidins, it did glucosylate flavanones and flavones in addition to retaining activity with flavonols. HPLC was used for product identification and showed mutant P145T glucosylated naringenin at the 7-OH position forming naringenin-7-O-glucoside and flavonols at the 3-OH position. Homology modeling and docking was done to predict the acceptor substrate recognition pattern and models were validated by experimental results. In other related work, a thrombin cleavage site was inserted into wild type Cp3GT and recombinant P145T enzyme between the enzyme and the C-myc tags in order to be able to cleave off tags. This provides the tool needed for future efforts to crystallize these proteins for structural determination. |
author |
Kandel, Sangam, Mr |
author_facet |
Kandel, Sangam, Mr |
author_sort |
Kandel, Sangam, Mr |
title |
Structural and Functional Analysis of Grapefruit Flavonol-Specific-3-O-GT Mutant P145T |
title_short |
Structural and Functional Analysis of Grapefruit Flavonol-Specific-3-O-GT Mutant P145T |
title_full |
Structural and Functional Analysis of Grapefruit Flavonol-Specific-3-O-GT Mutant P145T |
title_fullStr |
Structural and Functional Analysis of Grapefruit Flavonol-Specific-3-O-GT Mutant P145T |
title_full_unstemmed |
Structural and Functional Analysis of Grapefruit Flavonol-Specific-3-O-GT Mutant P145T |
title_sort |
structural and functional analysis of grapefruit flavonol-specific-3-o-gt mutant p145t |
publisher |
Digital Commons @ East Tennessee State University |
publishDate |
2016 |
url |
https://dc.etsu.edu/etd/3150 https://dc.etsu.edu/cgi/viewcontent.cgi?article=4560&context=etd |
work_keys_str_mv |
AT kandelsangammr structuralandfunctionalanalysisofgrapefruitflavonolspecific3ogtmutantp145t |
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1719189328197844992 |