The attachment of sugars to naturally-occurring and/or synthetic small molecules can dramatically influence the corresponding mechanism, pharmacodynamics, pharmacokinetics and even patent life of the parental structure. Yet, due to the technical challenges associated with conventional glycosylation strategies, the application of glycosylation in the context of drug discovery and/or development remains underexplored. An overarching aim of this program has been to develop simple and user-friendly chemoenzymatic methods for the differential glycosylation of target scaffolds to ultimately enable such drug discovery/development exploration. Toward this goal, the first phase of this study (years 1-5) led to the key proof of concept for chemoenzymatic glycorandomization (a one pot, three enzyme strategy capable of activating and attaching free monosaccharides to complex natural product scaffolds) and also provided fundamental information regarding two critical, but poorly understood, enzyme classes (anomeric sugar kinases and sugar-1-phosphate nucleotidylyltransferases). The second phase of this project (years 6-10) dramatically expanded the attempted application of this first generation system toward a range of diverse scaffolds, leading to success in many cases but also exposing key limitations of the platform. Importantly, the work conducted during the second phase also led to fundamental new knowledge regarding glycosyltransferase (GT)-catalyzed reactions that serves as the basis from which to launch the third phase of this study. Specifically, the proposed third phase of this study (years 11-15) takes advantage of our recent abilities to evolve highly permissive GTs and also drive GT-catalyzed reactions in reverse to enable next generation single enzyme or dual enzyme transglycosylation strategies for differential glycosylation of an array of structurally-diverse scaffolds (including natural product-based or synthetic, glycosylated or non-glycosylated, parental scaffolds). We propose to take full advantage of the current state of the art to: i) narrow the gaps of knowledge in understanding functional GT structure-activity-relationships;ii) specifically develop a range o catalysts for the production of novel sugar nucleotides (anticipated to be of broad use to the glycobiology community);iii) develop a range of catalysts for the differential glycosylation of a key set of structurally diverse anti-infective and anticancer scaffolds;and iv)exploit the corresponding differentially glycosylated scaffolds as a novel source for the discovery of new antiinfective and anticancer leads.

Public Health Relevance

We propose to develop simple and user-friendly chemoenzymatic methods for the differential glycosylation of complex target scaffolds to enable glycoconjugation as a tool for drug discovery and development. The models selected as part of the current study are anticipated to hold particularly high potential for the discovery of new antibacterial, antiviral and anticancer leads.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37AI052218-13
Application #
8663166
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Tseng, Christopher K
Project Start
2002-06-03
Project End
2017-05-31
Budget Start
2014-06-01
Budget End
2015-05-31
Support Year
13
Fiscal Year
2014
Total Cost
Indirect Cost
Name
University of Kentucky
Department
Pharmacology
Type
Schools of Pharmacy
DUNS #
City
Lexington
State
KY
Country
United States
Zip Code
40506
Huber, Tyler D; Wang, Fengbin; Singh, Shanteri et al. (2016) Functional AdoMet Isosteres Resistant to Classical AdoMet Degradation Pathways. ACS Chem Biol 11:2484-91
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Singh, Shanteri; Michalska, Karolina; Bigelow, Lance et al. (2015) Structural Characterization of CalS8, a TDP-α-D-Glucose Dehydrogenase Involved in Calicheamicin Aminodideoxypentose Biosynthesis. J Biol Chem 290:26249-58
Peltier-Pain, Pauline; Singh, Shanteri; Thorson, Jon S (2015) Characterization of Early Enzymes Involved in TDP-Aminodideoxypentose Biosynthesis en Route to Indolocarbazole AT2433. Chembiochem 16:2141-6
Elshahawi, Sherif I; Shaaban, Khaled A; Kharel, Madan K et al. (2015) A comprehensive review of glycosylated bacterial natural products. Chem Soc Rev 44:7591-697
Singh, Shanteri; Peltier-Pain, Pauline; Tonelli, Marco et al. (2014) A general NMR-based strategy for the in situ characterization of sugar-nucleotide-dependent biosynthetic pathways. Org Lett 16:3220-3
Zhang, Jianjun; Singh, Shanteri; Hughes, Ryan R et al. (2014) A simple strategy for glycosyltransferase-catalyzed aminosugar nucleotide synthesis. Chembiochem 15:647-52

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