Glycans decorating N- and O-linked glycoproteins, which constitute critical elements of the cell-surface landscape of many Gram-negative pathogens, integrate a variety of prokaryote-specific carbohydrates including di-N-acetyl bacillosamine (diNAcBac) and pseudaminic acid (Pse). There is growing genetic and biological evidence that modified saccharides, such as diNAcBac and Pse, are essential elements in the prokaryotic glycoconjugate repertoire and that cell surface glycoproteins that feature these sugars, serve as virulence factors, which mediate pathogen-host interactions and contribute to the severity of microbial infections. Previous studies have highlighted the fact that there is a striking diversity of monosaccharides in prokaryotes, relative to eukaryotes, however, there is a major unmet need for synthetic small molecule inhibitors that can be used as selective tools to acutely perturb their biosynthesis to understand the associations between modified sugars and bacterial pathogenicity. The proposed research involves fragment-based inhibitor design and structure-guided ligand optimization approaches together with incisive in vitro and in vivo analyses in the development of inhibitors of amino sugar acetyl transferases that catalyze key steps in the biosynthesis of UDP-diNAcBac and CMP-Pse. DiNAcBac and Pse are particularly prevalent microbial carbohydrates, which feature in the N- and O-linked glycoproteins of C. jejuni, A. baumannii and N. gonorrhoeae. These microbial pathogens are the targets of this research due to the established connections between protein glycosylation and virulence. We propose that small molecule inhibitors that acutely inhibit essential early steps in glycoprotein biosynthesis will allow for temporal control of glycoprotein biosynthesis that is not feasible with genetic approaches alone. Such inhibitors will be valuable new chemical tools that can provide insight into the effects of acutely inhibiting glycoprotein biosynthesis on motility, adherence and invasion in the native pathogen and in a pathogen/host context. The availability of inhibitors with appropriate biological properties will also validate the essentiality of carbohydrate modifications on microbial virulence in microorganisms (C. jejuni and N. gonorrhoeae) where genetic phenotyping and animal studies have provided clear evidence of the connections between glycosylation and virulence. This research will form a foundation for the application of similar strategic approaches with other pathogens that threaten human health where bioanalytical and bioinformatics approaches have been employed to predict the existence of glycoproteins that include highly modified carbohydrate building blocks. Ultimately we will address the central hypothesis that the enzymes that catalyze formation of unusual microbe-specific carbohydrate building blocks represent an "Achilles'heel" that can be exploited in the development of agents that can attenuate the virulence of serious human pathogens, which can be exploited in the battle against infectious diseases.

Public Health Relevance

Microbial pathogens decorate their cell surfaces with glycoprotein virulence factors, which promote access to and attack on human cells. Recent research has shown that these glycoproteins include a dazzling array of unusual pathogen-specific carbohydrate building blocks. Our research supports studies to develop selective chemical tools to elucidate the roles of modified carbohydrates in pathogenesis and addresses the central hypothesis that the biosynthetic pathways that lead to these building blocks, represent an Achilles'heel that can be exploited in the development of agents that can attenuate the virulence of serious human pathogens.

Agency
National Institute of Health (NIH)
Type
Research Project (R01)
Project #
2R01GM097241-03
Application #
8757021
Study Section
Macromolecular Structure and Function A Study Section (MSFA)
Program Officer
Marino, Pamela
Project Start
Project End
Budget Start
Budget End
Support Year
3
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Massachusetts Institute of Technology
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
City
Cambridge
State
MA
Country
United States
Zip Code
02142