[R01] Importance of Post-translational modifications in Streptococcus mutans pathophysiology
Ente: National Institute of Dental and Craniofacial Research
Scadenza: 2030-05-31
Importo max: 379.791 EUR
Paese: US
Descrizione
Abstract
Dental caries affects 2.5 billion people worldwide. Streptococcus mutans is an important pathogen in dental
caries due to its biofilm lifestyle, and acidogenic and aciduric nature. In addition, expression of the collagen
binding protein Cnm, present in 20% of the strains, by S. mutans is intimately associated with severe caries,
systemic infections and auto-immune disorders. Our mass-spectrometry studies have demonstrated that the
threonine-rich repeat (TRR) domain of S. mutans Cnm, with >70 threonine residues, undergoes extensive O-
glycosylation with N-acetylhexosamines (HexNAc2). We also found that cnm is co-transcribed with a
downstream gene encoding a putative GT-A type glycosyltransferase, named pgfS (protein glycosyltransferase
of streptococci). We identified three genes immediately downstream of pgfS, namely pgfM1, pgfE and pgfM2,
that also contribute to Cnm modification. Importantly, the pgf genes are part of the S. mutans core genome and
our findings suggest that we have uncovered an O-glycosylation pathway dedicated to modifying multiple
carbohydrate binding adhesins. Surprisingly, we also detected phosphate modifications in the TRR domain of
Cnm in pgfS and pgfM2 mutants, and also in low levels in wildtype Cnm. Based on structural homology
comparisons and extensive preliminary data, we hypothesize that post-translational modifications
significantly contribute to the complexity of S. mutans proteome, playing an important role in S. mutans
pathobiology. Specifically, O-linked protein glycosylation of surface proteins in S. mutans proceeds
through a series of lipid-linked glycan intermediates and protein kinase modifications. This is important
since this same lipid is essential for the biosynthesis of both peptidoglycan and the surface associated rhamnose-
glucose polysaccharide (RGP) and therefore regulatory mechanisms must be in place to control its distribution
among all three pathways (adhesin glycosylation, and peptidoglycan and RGP biosynthesis). In aim 1 we
propose to Characterize the enzymatic function of PgfS, PgfM1 and PgfM2 and localize the cellular
orientation of the active sites. In aim 2, we will determine O-glycosylation of other glycan binding surface
proteins by the Pgf machinery. Lastly, in aim 3 we will test the impact of the interplay between
glycosylation and phosphorylation on S. mutans fitness and pathophysiology. Our goal is to fill major gaps
in knowledge on how cell wall perturbations through anchoring of surface glycoproteins and RGP impact S.
mutans pathophysiology. Moreover, in this basic science proposal, using S. mutans as a model organism, our
findings will likely reveal novel mechanisms of post-translational modifications that could have broader impact
on our understanding of cell envelope homeostasis in Gram-positive bacteria.
Istituzione: UNIVERSITY OF FLORIDA
PI: Jacqueline Abranches, Christine M. Szymanski
Progetto: 5R01DE022559-13
Settori: National Institute of Dental and Craniofacial Research
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