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Enzyme Activity Measurement for Glycosyl, Hexosyl, and Pentosyl Transferases

Creative Enzymes provides accurate and reproducible enzyme activity assays for glycosyltransferases, including hexosyltransferases and pentosyltransferases. Our services do not stand out among all service suppliers only because of the difficulty of conducting the assays, but also because of the excellent quality of test results and consistency in data collection. Creative Enzymes has been the expertise in glycosyltransferase activity quantification and will continue delivering satisfactory results.

Glycosyltransferases (GTs, GTFs) are a group of enzymes, designated as EC 2.4, that catalyze the formation of the glycosidic linkage to form a glycoside. They transfer the glycosyl group from an activated nucleotide sugar, known as the glycosyl donor and often a sugar phosphate, to a nucleophilic glycosyl acceptor molecule. The nucleophilic group of the acceptor could be an oxygen- carbon-, nitrogen-, or sulfur-based group, producing the corresponding O-, C-, N-, or S-glycosides, which may be part of a monosaccharide, oligosaccharide, or polysaccharide. Most glycosyltransferases rely on metal ions to catalyze the reaction, since the phosphate group of the donor are usually coordinated by divalent cations such as manganese. However, metal independent enzymes exist.

Based on the stereochemistry of the product, EC 2.4 enzymes can be divided into inverting glycosyltransferases and retaining glycosyltransferases. Indicated by their names, inverting glycosyltransferases give products with a direction of the glycosidic bond opposite to that in the substrates, while the product of a retaining glycosyltransferase presents a configuration of the anomic center inverted from the substrate. Together the two enzymes demonstrate the variability of the natural biosynthesis which is hardly achieved by chemical synthesis. Alternatively, glycosyltransferases can also be categorized into about 100 GT families based on the result of alignment of the amino acid sequence. Each GT family contains enzymes that are related by sequence, corollary, and folding. Therefore, the catalytic mechanism is mostly conserved within each family, which then allows informative predictions to be made for GTs that are less studied.

Glycosyltransferases showed huge potential mainly in biomedical applications and synthesis of various glycosides and glycoconjugates, along with less common but critical products for dietary nutrition and environmental care. As mentioned above, GTs are highly efficient in making region- and stereo-specific glycosides, and thus are used in the both targeted synthesis of specific glycoconjugates as well as the construction of differentially glycosylated libraries. On the other side, because some oligo- and poly-saccharides are important to post-translational modifications of various proteins and biosynthesis of cell membranes, GTs are also popular targets as potential treatment of many diseases. For example, ethambutol is an inhibitor of mycobacterial arabinotransferases and used for the treatment of tuberculosis; caspofungin has been developed from the echinocandins and is an antifungal agent; moenomycin, an inhibitor of peptidoglycan glycosyltransferases, is used in animal feed as a growth promoter.

Overall, GTs are more and more studied over recent years targeting higher catalytic efficiency and more stable performance. However, as the first step of enzyme characterization, the activity assays for GTs are difficult to carry out, generally due to low availability of the substrates and lack of sensitive detection methods. To solve the challenge, Creative Enzymes has developed various tests that are able to accurately monitor enzyme activity, which include chromatographic assays and coupled enzymatic assays. A majority of these assays are technically challenging and can be properly performed only by few companies. Creative Enzymes is proud to present these tests to customers from all industries. With the support of a strong team of scientists and the advantage of high-end analytical equipment, we are recognized with consistent, high quality of the activity measurement. We encourage each client to share as many details of the analytical request as possible to ensure selection and optimization of the most suitable assays.

Enzyme Activity Measurement for Glycosyl, Hexosyl, and Pentosyl Transferases Figure: Unrooted phylogenetic tree of β-Kdo glycosyltransferases, suggested by multiple sequence alignment.
Reference: Ovchinnikova, O. G.; Mallette, E.; Koizumi, A.; Lowary, T. L.; Kimber, M. S.; Whitfield, C. Proc. Natl. Acad. Sci. 2016.

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