Unconventional N-Linked Glycosylation Promotes Trimeric Autotransporter Function in <named-content content-type="genus-species">Kingella kingae</named-content> and <named-content content-type="genus-species">Aggregatibacter aphrophilus</named-content>

ABSTRACT Glycosylation is a widespread mechanism employed by both eukaryotes and bacteria to increase the functional diversity of their proteomes. The nontypeable Haemophilus influenzae glycosyltransferase HMW1C mediates unconventional N-linked glycosylation of the adhesive protein HMW1, which is en...

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Autores principales:	Katherine A. Rempe, Lynn A. Spruce, Eric A. Porsch, Steven H. Seeholzer, Niels Nørskov-Lauritsen, Joseph W. St. Geme
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2015
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/bc9e5199e0834bdf9e35ce69e41550e8
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spelling	oai:doaj.org-article:bc9e5199e0834bdf9e35ce69e41550e82021-11-15T15:41:25ZUnconventional N-Linked Glycosylation Promotes Trimeric Autotransporter Function in <named-content content-type="genus-species">Kingella kingae</named-content> and <named-content content-type="genus-species">Aggregatibacter aphrophilus</named-content>10.1128/mBio.01206-152150-7511https://doaj.org/article/bc9e5199e0834bdf9e35ce69e41550e82015-09-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01206-15https://doaj.org/toc/2150-7511ABSTRACT Glycosylation is a widespread mechanism employed by both eukaryotes and bacteria to increase the functional diversity of their proteomes. The nontypeable Haemophilus influenzae glycosyltransferase HMW1C mediates unconventional N-linked glycosylation of the adhesive protein HMW1, which is encoded in a two-partner secretion system gene cluster that also encodes HMW1C. In this system, HMW1 is modified in the cytoplasm by sequential transfer of hexose residues. In the present study, we examined Kingella kingae and Aggregatibacter aphrophilus homologues of HMW1C that are not encoded near a gene encoding an obvious acceptor protein. We found both homologues to be functional glycosyltransferases and identified their substrates as the K. kingae Knh and the A. aphrophilus EmaA trimeric autotransporter proteins. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis revealed multiple sites of N-linked glycosylation on Knh and EmaA. Without glycosylation, Knh and EmaA failed to facilitate wild-type levels of bacterial autoaggregation or adherence to human epithelial cells, establishing that glycosylation is essential for proper protein function. IMPORTANCE This work emphasizes the importance of glycosylation for proper function of bacterial proteins. Here we show that the Kingella kingae Knh and the Aggregatibacter aphrophilus EmaA trimeric autotransporter proteins are N-glycosylated by novel homologues of the Haemophilus influenzae HMW1C glycosyltransferase, highlighting the first examples of trimeric autotransporters that are modified by HMW1C-like enzymes. In the absence of glycosylation, Knh and EmaA lack adhesive activity. This work has relevance to our understanding of bacterial pathogenicity and expression of potential vaccine antigens.Katherine A. RempeLynn A. SpruceEric A. PorschSteven H. SeeholzerNiels Nørskov-LauritsenJoseph W. St. GemeAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 6, Iss 4 (2015)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Katherine A. Rempe Lynn A. Spruce Eric A. Porsch Steven H. Seeholzer Niels Nørskov-Lauritsen Joseph W. St. Geme Unconventional N-Linked Glycosylation Promotes Trimeric Autotransporter Function in <named-content content-type="genus-species">Kingella kingae</named-content> and <named-content content-type="genus-species">Aggregatibacter aphrophilus</named-content>
description	ABSTRACT Glycosylation is a widespread mechanism employed by both eukaryotes and bacteria to increase the functional diversity of their proteomes. The nontypeable Haemophilus influenzae glycosyltransferase HMW1C mediates unconventional N-linked glycosylation of the adhesive protein HMW1, which is encoded in a two-partner secretion system gene cluster that also encodes HMW1C. In this system, HMW1 is modified in the cytoplasm by sequential transfer of hexose residues. In the present study, we examined Kingella kingae and Aggregatibacter aphrophilus homologues of HMW1C that are not encoded near a gene encoding an obvious acceptor protein. We found both homologues to be functional glycosyltransferases and identified their substrates as the K. kingae Knh and the A. aphrophilus EmaA trimeric autotransporter proteins. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis revealed multiple sites of N-linked glycosylation on Knh and EmaA. Without glycosylation, Knh and EmaA failed to facilitate wild-type levels of bacterial autoaggregation or adherence to human epithelial cells, establishing that glycosylation is essential for proper protein function. IMPORTANCE This work emphasizes the importance of glycosylation for proper function of bacterial proteins. Here we show that the Kingella kingae Knh and the Aggregatibacter aphrophilus EmaA trimeric autotransporter proteins are N-glycosylated by novel homologues of the Haemophilus influenzae HMW1C glycosyltransferase, highlighting the first examples of trimeric autotransporters that are modified by HMW1C-like enzymes. In the absence of glycosylation, Knh and EmaA lack adhesive activity. This work has relevance to our understanding of bacterial pathogenicity and expression of potential vaccine antigens.
format	article
author	Katherine A. Rempe Lynn A. Spruce Eric A. Porsch Steven H. Seeholzer Niels Nørskov-Lauritsen Joseph W. St. Geme
author_facet	Katherine A. Rempe Lynn A. Spruce Eric A. Porsch Steven H. Seeholzer Niels Nørskov-Lauritsen Joseph W. St. Geme
author_sort	Katherine A. Rempe
title	Unconventional N-Linked Glycosylation Promotes Trimeric Autotransporter Function in <named-content content-type="genus-species">Kingella kingae</named-content> and <named-content content-type="genus-species">Aggregatibacter aphrophilus</named-content>
title_short	Unconventional N-Linked Glycosylation Promotes Trimeric Autotransporter Function in <named-content content-type="genus-species">Kingella kingae</named-content> and <named-content content-type="genus-species">Aggregatibacter aphrophilus</named-content>
title_full	Unconventional N-Linked Glycosylation Promotes Trimeric Autotransporter Function in <named-content content-type="genus-species">Kingella kingae</named-content> and <named-content content-type="genus-species">Aggregatibacter aphrophilus</named-content>
title_fullStr	Unconventional N-Linked Glycosylation Promotes Trimeric Autotransporter Function in <named-content content-type="genus-species">Kingella kingae</named-content> and <named-content content-type="genus-species">Aggregatibacter aphrophilus</named-content>
title_full_unstemmed	Unconventional N-Linked Glycosylation Promotes Trimeric Autotransporter Function in <named-content content-type="genus-species">Kingella kingae</named-content> and <named-content content-type="genus-species">Aggregatibacter aphrophilus</named-content>
title_sort	unconventional n-linked glycosylation promotes trimeric autotransporter function in <named-content content-type="genus-species">kingella kingae</named-content> and <named-content content-type="genus-species">aggregatibacter aphrophilus</named-content>
publisher	American Society for Microbiology
publishDate	2015
url	https://doaj.org/article/bc9e5199e0834bdf9e35ce69e41550e8
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Unconventional N-Linked Glycosylation Promotes Trimeric Autotransporter Function in <named-content content-type="genus-species">Kingella kingae</named-content> and <named-content content-type="genus-species">Aggregatibacter aphrophilus</named-content>

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