Selection against glycosylation sites in potential target proteins of the general HMWC N-glycosyltransferase in Haemophilus influenzae

Gawthorne, Jayde A., Tan, Nikki Y., Bailey, Ulla-Maja, Davis, Margaret R., Wong, Linette W., Naidu, Ranjitha, Fox, Kate L., Jennings, Michael P. and Schulz, Benjamin L. (2014) Selection against glycosylation sites in potential target proteins of the general HMWC N-glycosyltransferase in Haemophilus influenzae. Biochemical and Biophysical Research Communications, 445 3: 633-638. doi:10.1016/j.bbrc.2014.02.044


Author Gawthorne, Jayde A.
Tan, Nikki Y.
Bailey, Ulla-Maja
Davis, Margaret R.
Wong, Linette W.
Naidu, Ranjitha
Fox, Kate L.
Jennings, Michael P.
Schulz, Benjamin L.
Title Selection against glycosylation sites in potential target proteins of the general HMWC N-glycosyltransferase in Haemophilus influenzae
Formatted title
Selection against glycosylation sites in potential target proteins of the general HMWC N-glycosyltransferase in Haemophilus influenzae
Journal name Biochemical and Biophysical Research Communications   Check publisher's open access policy
ISSN 0006-291X
1090-2104
Publication date 2014-02-21
Sub-type Article (original research)
DOI 10.1016/j.bbrc.2014.02.044
Volume 445
Issue 3
Start page 633
End page 638
Total pages 6
Place of publication Maryland Heights, MO, United States
Publisher Elsevier
Collection year 2015
Language eng
Formatted abstract
The HMWABC system of non-typeable Haemophilus influenzae (NTHi) encodes the HMWA adhesin glycoprotein, which is glycosylated by the HMWC glycosyltransferase. HMWC is a cytoplasmic N-glycosyltransferase, homologues of which are widespread in the Pasteurellaceae. We developed an assay for nonbiased detection of glycoproteins in NTHi based on metabolic engineering of the Leloir pathway and growth in media containing radiolabelled monosaccharides. The only glycoprotein identified in NTHi by this assay was HMWA. However, glycoproteomic analyses ex vivo in Escherichia coli showed that HMWC of NTHi was a general glycosyltransferase capable of glycosylating selected asparagines in proteins other than its HMWA substrate, including Asn78 in E. coli 30S ribosomal protein S5. The equivalent residue in S5 homologues in H. influenzae or other sequenced Pasteurellaceae genomes is not asparagine, and these organisms also showed significantly fewer than expected potential sites of glycosylation in general. Expression of active HMWC in E. coli resulted in growth inhibition compared with expression of inactive enzyme, consistent with glycosylation by HMWC detrimentally affecting the function of some E. coli proteins. Together, this supports the presence of a selective pressure in the Pasteurellaceae against glycosylation sites that would be modified by the general N-glycosyltransferase activity of HMWC.
Keyword Glycobiology
Metabolic engineering
Mass spectrometry
Proteomics
Protein evolution
Leloir pathway
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2015 Collection
School of Chemistry and Molecular Biosciences
 
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Created: Fri, 28 Feb 2014, 13:06:45 EST by Mrs Louise Nimwegen on behalf of School of Chemistry & Molecular Biosciences