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Identification of bacterial protein O-oligosaccharyltransferases and their glycoprotein substrates.

Schulz BL, Jen FE, Power PM, Jones CE, Fox KL, Ku SC, Blanchfield JT, Jennings MP - PLoS ONE (2013)

Bottom Line: We show that in the general glycosylation system of N. meningitidis, efficient glycosylation of additional protein substrates requires local structural similarity to the pilin acceptor site.For some Neisserial PglL substrates identified by sensitive analytical approaches, only a small fraction of the total protein pool is modified in the native organism, whereas others are completely glycosylated.Our results show that bacterial protein O-glycosylation is common, and that substrate selection in the general Neisserial system is dominated by recognition of structural homology.

View Article: PubMed Central - PubMed

Affiliation: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Queensland, Australia.

ABSTRACT
O-glycosylation of proteins in Neisseria meningitidis is catalyzed by PglL, which belongs to a protein family including WaaL O-antigen ligases. We developed two hidden Markov models that identify 31 novel candidate PglL homologs in diverse bacterial species, and describe several conserved sequence and structural features. Most of these genes are adjacent to possible novel target proteins for glycosylation. We show that in the general glycosylation system of N. meningitidis, efficient glycosylation of additional protein substrates requires local structural similarity to the pilin acceptor site. For some Neisserial PglL substrates identified by sensitive analytical approaches, only a small fraction of the total protein pool is modified in the native organism, whereas others are completely glycosylated. Our results show that bacterial protein O-glycosylation is common, and that substrate selection in the general Neisserial system is dominated by recognition of structural homology.

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N.meningitidis PglL topology and conservation.(A) Transmembrane profile of N. meningitidis PglL with the regions identified by the PglL_A, PglL_B and Wzy_C hidden Markov models indicated by red, green and blue lines and highly conserved amino acids coloured in red and orange. (B) The PglL protein Phobius transmembrane helix prediction with predicted transmembrane regions represented by dashed-lines. (C) CLUSTALX plot of sequence conservation of CLUSTALW alignment of the putative PglL proteins. (D) Regions identified by the PglL_A, PglL_B and Wzy_C hidden Markov models indicated by red, green and blue boxes respectively.
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pone-0062768-g001: N.meningitidis PglL topology and conservation.(A) Transmembrane profile of N. meningitidis PglL with the regions identified by the PglL_A, PglL_B and Wzy_C hidden Markov models indicated by red, green and blue lines and highly conserved amino acids coloured in red and orange. (B) The PglL protein Phobius transmembrane helix prediction with predicted transmembrane regions represented by dashed-lines. (C) CLUSTALX plot of sequence conservation of CLUSTALW alignment of the putative PglL proteins. (D) Regions identified by the PglL_A, PglL_B and Wzy_C hidden Markov models indicated by red, green and blue boxes respectively.

Mentions: To identify PglL homologs in bacterial genomes we developed a hidden Markov model (HMM) that would resolve the subset of PglL protein O-OTases from the wider PFAM PF04932, which contains both WaaL O-antigen ligases and PglL proteins. This family of enzymes has low overall amino acid similarity but contains a small region of conservation that is the basis for PFAM PF04932. To identify sequence features which accounted for the protein acceptor substrate specificity of PglL, we performed multi-sequence alignments of protein sequences of close homologues of PglL and used conserved features not present in WaaL to create two HMMs, pglL_A and pglL_B (Fig. 1). HMM pgl_A has been submitted to the Pfam database with accession number PF15864. These HMMs did not identify well-characterized WaaL proteins from enteric organisms, suggesting that they may be useful for the identification of PglL candidates in wider searches.


Identification of bacterial protein O-oligosaccharyltransferases and their glycoprotein substrates.

Schulz BL, Jen FE, Power PM, Jones CE, Fox KL, Ku SC, Blanchfield JT, Jennings MP - PLoS ONE (2013)

N.meningitidis PglL topology and conservation.(A) Transmembrane profile of N. meningitidis PglL with the regions identified by the PglL_A, PglL_B and Wzy_C hidden Markov models indicated by red, green and blue lines and highly conserved amino acids coloured in red and orange. (B) The PglL protein Phobius transmembrane helix prediction with predicted transmembrane regions represented by dashed-lines. (C) CLUSTALX plot of sequence conservation of CLUSTALW alignment of the putative PglL proteins. (D) Regions identified by the PglL_A, PglL_B and Wzy_C hidden Markov models indicated by red, green and blue boxes respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3643930&req=5

pone-0062768-g001: N.meningitidis PglL topology and conservation.(A) Transmembrane profile of N. meningitidis PglL with the regions identified by the PglL_A, PglL_B and Wzy_C hidden Markov models indicated by red, green and blue lines and highly conserved amino acids coloured in red and orange. (B) The PglL protein Phobius transmembrane helix prediction with predicted transmembrane regions represented by dashed-lines. (C) CLUSTALX plot of sequence conservation of CLUSTALW alignment of the putative PglL proteins. (D) Regions identified by the PglL_A, PglL_B and Wzy_C hidden Markov models indicated by red, green and blue boxes respectively.
Mentions: To identify PglL homologs in bacterial genomes we developed a hidden Markov model (HMM) that would resolve the subset of PglL protein O-OTases from the wider PFAM PF04932, which contains both WaaL O-antigen ligases and PglL proteins. This family of enzymes has low overall amino acid similarity but contains a small region of conservation that is the basis for PFAM PF04932. To identify sequence features which accounted for the protein acceptor substrate specificity of PglL, we performed multi-sequence alignments of protein sequences of close homologues of PglL and used conserved features not present in WaaL to create two HMMs, pglL_A and pglL_B (Fig. 1). HMM pgl_A has been submitted to the Pfam database with accession number PF15864. These HMMs did not identify well-characterized WaaL proteins from enteric organisms, suggesting that they may be useful for the identification of PglL candidates in wider searches.

Bottom Line: We show that in the general glycosylation system of N. meningitidis, efficient glycosylation of additional protein substrates requires local structural similarity to the pilin acceptor site.For some Neisserial PglL substrates identified by sensitive analytical approaches, only a small fraction of the total protein pool is modified in the native organism, whereas others are completely glycosylated.Our results show that bacterial protein O-glycosylation is common, and that substrate selection in the general Neisserial system is dominated by recognition of structural homology.

View Article: PubMed Central - PubMed

Affiliation: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Queensland, Australia.

ABSTRACT
O-glycosylation of proteins in Neisseria meningitidis is catalyzed by PglL, which belongs to a protein family including WaaL O-antigen ligases. We developed two hidden Markov models that identify 31 novel candidate PglL homologs in diverse bacterial species, and describe several conserved sequence and structural features. Most of these genes are adjacent to possible novel target proteins for glycosylation. We show that in the general glycosylation system of N. meningitidis, efficient glycosylation of additional protein substrates requires local structural similarity to the pilin acceptor site. For some Neisserial PglL substrates identified by sensitive analytical approaches, only a small fraction of the total protein pool is modified in the native organism, whereas others are completely glycosylated. Our results show that bacterial protein O-glycosylation is common, and that substrate selection in the general Neisserial system is dominated by recognition of structural homology.

Show MeSH