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Impact of carbohydrate binding modules on transglycosylation efficiency in the glycoside hydrolase family 5 proteins
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Kasture, Madhura. Impact of carbohydrate binding modules on transglycosylation efficiency in the glycoside hydrolase family 5 proteins. Retrieved from https://doi.org/doi:10.7282/t3-c78h-4e12

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TitleImpact of carbohydrate binding modules on transglycosylation efficiency in the glycoside hydrolase family 5 proteins
NameKasture, Madhura (author); Chundawat, Shishir (chair); Rutgers University; School of Graduate Studies
Date Created2020
Other Date2020-01 (degree)
SubjectChemical and Biochemical Engineering, Glycosidases
Extent1 online resource (xiii, 80 pages) : illustrations
DescriptionOligosaccharides are significant in many biological processes such as modulating protein folding, cell interactions, and binding and are widely used in prebiotics and pharmaceutical industries. This rising popularity has accelerated the research for their sustainable source of generation. Enzymatic synthesis offers an alternative over standard chemical synthesis to produce these oligosaccharides with higher specificity and product purity. Transglycosylases are a class of glycoside hydrolases (GH) present in nature and are ideal biocatalysts for this process. Since these are few in number and limited in substrate specificity, over the past couple of years, extensive studies and advancements have been made to engineer glycoside hydrolases to reduce the hydrolytic activity of these enzymes to perform transglycosylation to produce oligosaccharides.

This thesis investigates the transglycosylation activity of glycoside hydrolases family five that has tethered carbohydrate-binding module (CBM) CBM3a. CBMs improve the catalytic efficiency by binding to the polysaccharide chain, making the substrate more accessible to the enzyme. The catalytic nucleophile of GH5 enzymes was substituted for alanine, glycine, and serine amino acids. Activity assay in the presence of an activated soluble sugar was performed to obtain a product profile for the enzymes. With the data from the activity assays, a kinetic model was developed. A structure-functional analysis was attempted to explain the results. It was observed that few enzymes with a mutated catalytic nucleophile show transglycosylation like activity in the presence of an appended CBM3a, suggesting that the presence of CBM3a affected the mechanism of the enzyme activity other than just increasing the local concentration of the substrate at the catalytic pocket. In summary, this study provides results that led further confidence in the active role of the CBM3a in the transglycosylation activity of the GH5 enzymes and understanding the mechanism.
NoteM.S.
NoteIncludes bibliographical references
Genretheses, ETD graduate
Persistent URLhttps://doi.org/doi:10.7282/t3-c78h-4e12
LanguageEnglish
CollectionSchool of Graduate Studies Electronic Theses and Dissertations
Organization NameRutgers, The State University of New Jersey
RightsThe author owns the copyright to this work.
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