N-Glycosylation of Campylobacter jejuni Surface Proteins Promotes Bacterial Fitness

Abofu Alemka,Christine M Szymanski,Jing Zheng,Harald Nothaft,B A Mccormick

doi:10.1128/iai.01370-12

Abofu Alemka, Christine M Szymanski + Show 3 more

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https://doi.org/10.1128/iai.01370-12

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Abstract

Campylobacter jejuni is the etiologic agent of human bacterial gastroenteritis worldwide. In contrast, despite heavy colonization, C. jejuni maintains a commensal mode of existence in chickens. The consumption of contaminated chicken products is thought to be the principal mode of C. jejuni transmission to the human population. C. jejuni harbors a system for N-linked protein glycosylation that has been well characterized and modifies more than 60 periplasmic and membrane-bound proteins. However, the precise role of this modification in the biology of C. jejuni remains unexplored. We hypothesized that the N-glycans protect C. jejuni surface proteins from the action of gut proteases. The C. jejuni pglB mutant, deficient in the expression of the oligosaccharyltransferase, exhibited reduced growth in medium supplemented with chicken cecal contents (CCC) compared with that of wild-type (WT) cells. Inactivation of the cecal proteases by heat treatment or with protease inhibitors completely restored bacterial viability and partially rescued bacterial growth. Physiological concentrations of trypsin, but not chymotrypsin, also reduced C. jejuni pglB mutant CFU. Live or dead staining indicated that CCC preferentially influenced C. jejuni growth as opposed to bacterial viability. We identified multiple chicken cecal proteases by mass fingerprinting. The use of protease inhibitors that target specific classes indicated that both metalloproteases and serine proteases were involved in the attenuated growth of the oligosaccharyltransferase mutant. In conclusion, protein N-linked glycosylation of surface proteins may enhance C. jejuni fitness by protecting bacterial proteins from cleavage due to gut proteases.

Highlights

Campylobacter jejuni is the etiologic agent of human bacterial gastroenteritis worldwide
We speculated that one of the reasons why C. jejuni N-glycosylates its surface proteins may be to protect itself from the harsh environment of the gut, which consists of several digestive proteases
Following an incubation period of 2 h in cecal contents (CCC), there was an approximately 5-fold reduction in the recovered CFU of the pglB mutant compared with that of the WT (Fig. 2A). This reduction was completely abolished by inactivation of chicken cecal proteases either by heat treatment or with protease inhibitors (Fig. 2A), indicating that the observed effect was due to proteases

Summary

Introduction

Campylobacter jejuni is the etiologic agent of human bacterial gastroenteritis worldwide. The C. jejuni pglB mutant, deficient in the expression of the oligosaccharyltransferase, exhibited reduced growth in medium supplemented with chicken cecal contents (CCC) compared with that of wild-type (WT) cells. The N-glycosylation pathway of C. jejuni has been extensively characterized [2, 8, 9] This pathway involves the transfer en bloc of a conserved heptasaccharide onto asparagine residues of the extended sequon D/E-X1-NX2-S/T (where X1 and X2 are any amino acid except proline) on nascent proteins in the periplasmic side of the inner membrane by the oligosaccharyltransferase (OST) PglB [2, 8, 10]. The precise role of this modification in the pathophysiology of C. jejuni remains unknown

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