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Substitute sweeteners: diverse bacterial oligosaccharyltransferases with unique N-glycosylation site preferences

The central enzyme in the Campylobacter jejuni asparagine-linked glycosylation pathway is the oligosaccharyltransferase (OST), PglB, which transfers preassembled glycans to specific asparagine residues in target proteins. While C. jejuni PglB (CjPglB) can transfer many diverse glycan structures, the...

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Autores principales: Ollis, Anne A., Chai, Yi, Natarajan, Aravind, Perregaux, Emily, Jaroentomeechai, Thapakorn, Guarino, Cassandra, Smith, Jessica, Zhang, Sheng, DeLisa, Matthew P.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4894442/
https://www.ncbi.nlm.nih.gov/pubmed/26482295
http://dx.doi.org/10.1038/srep15237
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author Ollis, Anne A.
Chai, Yi
Natarajan, Aravind
Perregaux, Emily
Jaroentomeechai, Thapakorn
Guarino, Cassandra
Smith, Jessica
Zhang, Sheng
DeLisa, Matthew P.
author_facet Ollis, Anne A.
Chai, Yi
Natarajan, Aravind
Perregaux, Emily
Jaroentomeechai, Thapakorn
Guarino, Cassandra
Smith, Jessica
Zhang, Sheng
DeLisa, Matthew P.
author_sort Ollis, Anne A.
collection PubMed
description The central enzyme in the Campylobacter jejuni asparagine-linked glycosylation pathway is the oligosaccharyltransferase (OST), PglB, which transfers preassembled glycans to specific asparagine residues in target proteins. While C. jejuni PglB (CjPglB) can transfer many diverse glycan structures, the acceptor sites that it recognizes are restricted predominantly to those having a negatively charged residue in the −2 position relative to the asparagine. Here, we investigated the acceptor-site preferences for 23 homologs with natural sequence variation compared to CjPglB. Using an ectopic trans-complementation assay for CjPglB function in glycosylation-competent Escherichia coli, we demonstrated in vivo activity for 16 of the candidate OSTs. Interestingly, the OSTs from Campylobacter coli, Campylobacter upsaliensis, Desulfovibrio desulfuricans, Desulfovibrio gigas, and Desulfovibrio vulgaris, exhibited significantly relaxed specificity towards the −2 position compared to CjPglB. These enzymes glycosylated minimal N-X-T motifs in multiple targets and each followed unique, as yet unknown, rules governing acceptor-site preferences. One notable example is D. gigas PglB, which was the only bacterial OST to glycosylate the Fc domain of human immunoglobulin G at its native ‘QYNST’ sequon. Overall, we find that a subset of bacterial OSTs follow their own rules for acceptor-site specificity, thereby expanding the glycoengineering toolbox with previously unavailable biocatalytic diversity.
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spelling pubmed-48944422016-06-10 Substitute sweeteners: diverse bacterial oligosaccharyltransferases with unique N-glycosylation site preferences Ollis, Anne A. Chai, Yi Natarajan, Aravind Perregaux, Emily Jaroentomeechai, Thapakorn Guarino, Cassandra Smith, Jessica Zhang, Sheng DeLisa, Matthew P. Sci Rep Article The central enzyme in the Campylobacter jejuni asparagine-linked glycosylation pathway is the oligosaccharyltransferase (OST), PglB, which transfers preassembled glycans to specific asparagine residues in target proteins. While C. jejuni PglB (CjPglB) can transfer many diverse glycan structures, the acceptor sites that it recognizes are restricted predominantly to those having a negatively charged residue in the −2 position relative to the asparagine. Here, we investigated the acceptor-site preferences for 23 homologs with natural sequence variation compared to CjPglB. Using an ectopic trans-complementation assay for CjPglB function in glycosylation-competent Escherichia coli, we demonstrated in vivo activity for 16 of the candidate OSTs. Interestingly, the OSTs from Campylobacter coli, Campylobacter upsaliensis, Desulfovibrio desulfuricans, Desulfovibrio gigas, and Desulfovibrio vulgaris, exhibited significantly relaxed specificity towards the −2 position compared to CjPglB. These enzymes glycosylated minimal N-X-T motifs in multiple targets and each followed unique, as yet unknown, rules governing acceptor-site preferences. One notable example is D. gigas PglB, which was the only bacterial OST to glycosylate the Fc domain of human immunoglobulin G at its native ‘QYNST’ sequon. Overall, we find that a subset of bacterial OSTs follow their own rules for acceptor-site specificity, thereby expanding the glycoengineering toolbox with previously unavailable biocatalytic diversity. Nature Publishing Group 2015-10-20 /pmc/articles/PMC4894442/ /pubmed/26482295 http://dx.doi.org/10.1038/srep15237 Text en Copyright © 2015, Macmillan Publishers Limited http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
spellingShingle Article
Ollis, Anne A.
Chai, Yi
Natarajan, Aravind
Perregaux, Emily
Jaroentomeechai, Thapakorn
Guarino, Cassandra
Smith, Jessica
Zhang, Sheng
DeLisa, Matthew P.
Substitute sweeteners: diverse bacterial oligosaccharyltransferases with unique N-glycosylation site preferences
title Substitute sweeteners: diverse bacterial oligosaccharyltransferases with unique N-glycosylation site preferences
title_full Substitute sweeteners: diverse bacterial oligosaccharyltransferases with unique N-glycosylation site preferences
title_fullStr Substitute sweeteners: diverse bacterial oligosaccharyltransferases with unique N-glycosylation site preferences
title_full_unstemmed Substitute sweeteners: diverse bacterial oligosaccharyltransferases with unique N-glycosylation site preferences
title_short Substitute sweeteners: diverse bacterial oligosaccharyltransferases with unique N-glycosylation site preferences
title_sort substitute sweeteners: diverse bacterial oligosaccharyltransferases with unique n-glycosylation site preferences
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4894442/
https://www.ncbi.nlm.nih.gov/pubmed/26482295
http://dx.doi.org/10.1038/srep15237
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