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Mass spectrometric analysis of the S‐layer proteins from Clostridium difficile demonstrates the absence of glycosylation
Author(s) -
Qazi Omar,
Hitchen Paul,
Tissot Bérangère,
Panico Maria,
Morris Howard R.,
Dell Anne,
Fairweather Neil
Publication year - 2009
Publication title -
journal of mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.475
H-Index - 121
eISSN - 1096-9888
pISSN - 1076-5174
DOI - 10.1002/jms.1514
Subject(s) - chemistry , glycosylation , s layer , glycan , molecular mass , clostridium difficile , matrix assisted laser desorption/ionization , cyanogen bromide , biochemistry , mass spectrometry , clostridium , microbiology and biotechnology , chromatography , desorption , bacteria , peptide sequence , glycoprotein , adsorption , gene , enzyme , organic chemistry , biology , genetics , antibiotics
Like many other bacterial cell surfaces, the cell wall of Clostridium difficile is also encapsulated by a proteinaceous paracrystalline layer, the surface (S)‐layer. In many bacterial species, the S‐layer proteins (SLPs) have been shown to be glycosylated, whereas in other species glycosylation is absent. Unusually, the S‐layer of C. difficile is composed of two distinct proteins, the high‐molecular weight (HMW) and low‐molecular‐weight (LMW) SLPs. Previous investigations have reported that one or both of these SLPs are glycosylated, though no definitive study has been conducted. We have used a variety of mass spectrometric approaches to analyse SLPs from a number of strains of C. difficile for the presence of associated glycans. Analysis of intact SLPs by matrix assisted laser desorption/ionisation time of flight (MALDI‐ToF) mass spectrometry demonstrated that the observed molecular masses matched the predicted masses of the LMW and HMW SLPs. Furthermore, analysis of Cyanogen bromide (CNBr) and tryptic peptides displayed no evidence of post‐translational modification. In the first in‐depth study of its kind, we unequivocally demonstrate that the S‐layer proteins from the C. difficile strains investigated are not glycosylated. Copyright © 2008 John Wiley & Sons, Ltd.