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Degree Name
MS (Master of Science)
Program
Biology
Date of Award
8-2015
Committee Chair or Co-Chairs
Cecilia A. McIntosh
Committee Members
Aruna Kilaru, Shivakumar P. Devaiah
Abstract
A flavonol specific glucosyltransferase cloned from Citrus paradisi has strict substrate and regio-specificity (Cp3OGT). The amino acid sequence of Cp3OGT was aligned with sequences of an anthocyanidin UDP- dependant glucosyltransferase (UGT) from Clitorea ternatea and a UGT from Vitis vinifera that can glucosylate both flavonols and anthocyanidins. Using homology modeling to identify candidate regions followed by site directed mutagenesis, three double mutations were constructed and biochemically characterized. S20G+T21S mutant protein retained activity with flavonols similar to the wildtype Cp3OGT but the mutant had optimum activity at 60°C and broadened substrate acceptance to include the flavanone naringenin. S290C+S319A mutant protein retained 40% activity with quercetin relative to WT, and had an optimum pH shift. H154Y+Q87I mutant protein was only 10% active with quercetin relative to WT. Docking analysis revealed that H154, Q87 and S20 could be involved in orienting the acceptor molecules within the acceptor binding site whereas S319 and S290 residues are involved in maintaining the active site conformation.
Document Type
Thesis - restricted
Recommended Citation
Sathanantham, Preethi, "Impact of Mutations of Targeted Serine, Histidine, and Glutamine Residues in Citrus paradisi Flavonol Specific Glucosyltransferase Activity" (2015). Electronic Theses and Dissertations. Paper 2560. https://dc.etsu.edu/etd/2560
Copyright
Copyright by the authors.