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O-Glycosylation Landscapes of SARS-CoV-2 Spike Proteins

The densely glycosylated spike (S) proteins that are highly exposed on the surface of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) facilitate viral attachment, entry, and membrane fusion. We have previously reported all the 22 N-glycosites and site-specific N-glycans in the S protein...

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Autores principales: Zhang, Yong, Zhao, Wanjun, Mao, Yonghong, Chen, Yaohui, Zheng, Shanshan, Cao, Wei, Zhu, Jingqiang, Hu, Liqiang, Gong, Meng, Cheng, Jingqiu, Yang, Hao
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8450404/
https://www.ncbi.nlm.nih.gov/pubmed/34552909
http://dx.doi.org/10.3389/fchem.2021.689521
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author Zhang, Yong
Zhao, Wanjun
Mao, Yonghong
Chen, Yaohui
Zheng, Shanshan
Cao, Wei
Zhu, Jingqiang
Hu, Liqiang
Gong, Meng
Cheng, Jingqiu
Yang, Hao
author_facet Zhang, Yong
Zhao, Wanjun
Mao, Yonghong
Chen, Yaohui
Zheng, Shanshan
Cao, Wei
Zhu, Jingqiang
Hu, Liqiang
Gong, Meng
Cheng, Jingqiu
Yang, Hao
author_sort Zhang, Yong
collection PubMed
description The densely glycosylated spike (S) proteins that are highly exposed on the surface of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) facilitate viral attachment, entry, and membrane fusion. We have previously reported all the 22 N-glycosites and site-specific N-glycans in the S protein protomer. Herein, we report the O-glycosylation landscapes of SARS-CoV-2 S proteins, which were characterized through high-resolution mass spectrometry. Following digestion with trypsin and trypsin/Glu-C, and de-N-glycosylation using PNGase F, we determined the GalNAc-type O-glycosylation pattern of S proteins, including O-glycosites and the six most common O-glycans occupying them, via Byonic identification and manual validation. Finally, 255 intact O-glycopeptides composed of 50 peptides sequences and 43 O-glycosites were discovered by higher energy collision-induced dissociation (HCD), and three O-glycosites were confidently identified by electron transfer/higher energy collision-induced dissociation (EThcD) in the insect cell-expressed S protein. Most glycosites were modified by non-sialylated O-glycans such as HexNAc(1) and HexNAc(1)Hex (1). In contrast, in the human cell-expressed S protein S1 subunit, 407 intact O-glycopeptides composed of 34 peptides sequences and 30 O-glycosites were discovered by HCD, and 11 O-glycosites were unambiguously assigned by EThcD. However, the measurement of O-glycosylation occupancy hasn’t been made. Most glycosites were modified by sialylated O-glycans such as HexNAc(1)Hex (1)NeuAc (1) and HexNAc(1)Hex (1)NeuAc (2). Our results reveal that the SARS-CoV-2 S protein is an O-glycoprotein; the O-glycosites and O-glycan compositions vary with the host cell type. These comprehensive O-glycosylation landscapes of the S protein are expected to provide novel insights into the viral binding mechanism and present a strategy for the development of vaccines and targeted drugs.
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spelling pubmed-84504042021-09-21 O-Glycosylation Landscapes of SARS-CoV-2 Spike Proteins Zhang, Yong Zhao, Wanjun Mao, Yonghong Chen, Yaohui Zheng, Shanshan Cao, Wei Zhu, Jingqiang Hu, Liqiang Gong, Meng Cheng, Jingqiu Yang, Hao Front Chem Chemistry The densely glycosylated spike (S) proteins that are highly exposed on the surface of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) facilitate viral attachment, entry, and membrane fusion. We have previously reported all the 22 N-glycosites and site-specific N-glycans in the S protein protomer. Herein, we report the O-glycosylation landscapes of SARS-CoV-2 S proteins, which were characterized through high-resolution mass spectrometry. Following digestion with trypsin and trypsin/Glu-C, and de-N-glycosylation using PNGase F, we determined the GalNAc-type O-glycosylation pattern of S proteins, including O-glycosites and the six most common O-glycans occupying them, via Byonic identification and manual validation. Finally, 255 intact O-glycopeptides composed of 50 peptides sequences and 43 O-glycosites were discovered by higher energy collision-induced dissociation (HCD), and three O-glycosites were confidently identified by electron transfer/higher energy collision-induced dissociation (EThcD) in the insect cell-expressed S protein. Most glycosites were modified by non-sialylated O-glycans such as HexNAc(1) and HexNAc(1)Hex (1). In contrast, in the human cell-expressed S protein S1 subunit, 407 intact O-glycopeptides composed of 34 peptides sequences and 30 O-glycosites were discovered by HCD, and 11 O-glycosites were unambiguously assigned by EThcD. However, the measurement of O-glycosylation occupancy hasn’t been made. Most glycosites were modified by sialylated O-glycans such as HexNAc(1)Hex (1)NeuAc (1) and HexNAc(1)Hex (1)NeuAc (2). Our results reveal that the SARS-CoV-2 S protein is an O-glycoprotein; the O-glycosites and O-glycan compositions vary with the host cell type. These comprehensive O-glycosylation landscapes of the S protein are expected to provide novel insights into the viral binding mechanism and present a strategy for the development of vaccines and targeted drugs. Frontiers Media S.A. 2021-09-06 /pmc/articles/PMC8450404/ /pubmed/34552909 http://dx.doi.org/10.3389/fchem.2021.689521 Text en Copyright © 2021 Zhang, Zhao, Mao, Chen, Zheng, Cao, Zhu, Hu, Gong, Cheng and Yang. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Chemistry
Zhang, Yong
Zhao, Wanjun
Mao, Yonghong
Chen, Yaohui
Zheng, Shanshan
Cao, Wei
Zhu, Jingqiang
Hu, Liqiang
Gong, Meng
Cheng, Jingqiu
Yang, Hao
O-Glycosylation Landscapes of SARS-CoV-2 Spike Proteins
title O-Glycosylation Landscapes of SARS-CoV-2 Spike Proteins
title_full O-Glycosylation Landscapes of SARS-CoV-2 Spike Proteins
title_fullStr O-Glycosylation Landscapes of SARS-CoV-2 Spike Proteins
title_full_unstemmed O-Glycosylation Landscapes of SARS-CoV-2 Spike Proteins
title_short O-Glycosylation Landscapes of SARS-CoV-2 Spike Proteins
title_sort o-glycosylation landscapes of sars-cov-2 spike proteins
topic Chemistry
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8450404/
https://www.ncbi.nlm.nih.gov/pubmed/34552909
http://dx.doi.org/10.3389/fchem.2021.689521
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