Structural and Quantitative Characterization of Mucin-Type <i>O</i>-Glycans and the Identification of <i>O</i>-Glycosylation Sites in Bovine Submaxillary Mucin

Bovine submaxillary mucin (BSM) is a gel-forming glycoprotein polymer, and Ser/Thr-linked glycans (<i>O</i>-glycans) are important in regulating BSM’s viscoelasticity and polymerization. However, details of <i>O</i>-glycosylation have not been reported. This study investigate...

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Main Authors: Jihye Kim, Changsoo Ryu, Jongkwan Ha, Junmyoung Lee, Donghwi Kim, Minkyoo Ji, Chi Soo Park, Jaeryong Lee, Dae Kyong Kim, Ha Hyung Kim
Format: Article
Language:English
Published: MDPI AG 2020-04-01
Series:Biomolecules
Subjects:
Online Access:https://www.mdpi.com/2218-273X/10/4/636
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spelling doaj-70b51f4820d74fb7a03a40ad4fbed6af2020-11-25T02:21:36ZengMDPI AGBiomolecules2218-273X2020-04-011063663610.3390/biom10040636Structural and Quantitative Characterization of Mucin-Type <i>O</i>-Glycans and the Identification of <i>O</i>-Glycosylation Sites in Bovine Submaxillary MucinJihye Kim0Changsoo Ryu1Jongkwan Ha2Junmyoung Lee3Donghwi Kim4Minkyoo Ji5Chi Soo Park6Jaeryong Lee7Dae Kyong Kim8Ha Hyung Kim9Biotherapeutics and Glycomics Laboratory, College of Pharmacy, Chung-Ang University, Seoul 06974, KoreaBiotherapeutics and Glycomics Laboratory, College of Pharmacy, Chung-Ang University, Seoul 06974, KoreaBiotherapeutics and Glycomics Laboratory, College of Pharmacy, Chung-Ang University, Seoul 06974, KoreaBiotherapeutics and Glycomics Laboratory, College of Pharmacy, Chung-Ang University, Seoul 06974, KoreaBiotherapeutics and Glycomics Laboratory, College of Pharmacy, Chung-Ang University, Seoul 06974, KoreaBiotherapeutics and Glycomics Laboratory, College of Pharmacy, Chung-Ang University, Seoul 06974, KoreaBiotherapeutics and Glycomics Laboratory, College of Pharmacy, Chung-Ang University, Seoul 06974, KoreaBiotherapeutics and Glycomics Laboratory, College of Pharmacy, Chung-Ang University, Seoul 06974, KoreaDepartment of Environmental & Health Chemistry, College of Pharmacy, Chung-Ang University, Seoul 06974, KoreaBiotherapeutics and Glycomics Laboratory, College of Pharmacy, Chung-Ang University, Seoul 06974, KoreaBovine submaxillary mucin (BSM) is a gel-forming glycoprotein polymer, and Ser/Thr-linked glycans (<i>O</i>-glycans) are important in regulating BSM’s viscoelasticity and polymerization. However, details of <i>O</i>-glycosylation have not been reported. This study investigates the structural and quantitative characteristics of <i>O</i>-glycans and identifies <i>O</i>-glycosylation sites in BSM using liquid chromatography–tandem mass spectrometry. The <i>O</i>-glycans (consisting of di- to octa-saccharides) and their quantities (%) relative to total <i>O</i>-glycans (100%; 1.1 pmol per 1 μg of BSM) were identified with 14 major (>1.0%), 12 minor (0.1%–1.0%), and eight trace (<0.1%) <i>O</i>-glycans, which were characterized based on their constituents (sialylation (14 <i>O</i>-glycans; 81.9%, sum of relative quantities of each glycan), non-sialylation (20; 18.1%), fucosylation (20; 17.5%), and terminal-galactosylation (6; 3.6%)) and six core structure types [Gal-GalNAc, Gal-(GlcNAc)GalNAc, GlcNAc-GalNAc, GlcNAc-(GlcNAc)GalNAc, and GalNAc-GalNAc]. <i>O</i>-glycosylation sites were identified using <i>O</i>-glycopeptides (bold underlined; <sub>56</sub><b>S</b>GE<b>T</b>R<b>TS</b>VI, <sub>259</sub><b>S</b>H<b>SSS</b>GR<b>S</b>R<b>T</b>I, <sub>272</sub>G<b>S</b>P<b>SS</b>V<b>SS</b>AEQI, <sub>307</sub>RP<b>S</b>YGAL, <sub>625</sub>Q<b>T</b>LGPL, <sub>728</sub><b>T</b>M<b>TT</b>R<b>TS</b>VVV, and <sub>1080</sub>RPEDN<b>T</b>AVA) obtained from proteolytic BSM; these sites are in the four domains of BSM. The gel-forming mucins share common domain structures and glycosylation patterns; these results could provide useful information on mucin-type <i>O</i>-glycans. This is the first study to characterize <i>O</i>-glycans and identify <i>O</i>-glycosylation sites in BSM.https://www.mdpi.com/2218-273X/10/4/636bovine submaxillary mucinmucin-type O-glycanO-glycosylation site
collection DOAJ
language English
format Article
sources DOAJ
author Jihye Kim
Changsoo Ryu
Jongkwan Ha
Junmyoung Lee
Donghwi Kim
Minkyoo Ji
Chi Soo Park
Jaeryong Lee
Dae Kyong Kim
Ha Hyung Kim
spellingShingle Jihye Kim
Changsoo Ryu
Jongkwan Ha
Junmyoung Lee
Donghwi Kim
Minkyoo Ji
Chi Soo Park
Jaeryong Lee
Dae Kyong Kim
Ha Hyung Kim
Structural and Quantitative Characterization of Mucin-Type <i>O</i>-Glycans and the Identification of <i>O</i>-Glycosylation Sites in Bovine Submaxillary Mucin
Biomolecules
bovine submaxillary mucin
mucin-type O-glycan
O-glycosylation site
author_facet Jihye Kim
Changsoo Ryu
Jongkwan Ha
Junmyoung Lee
Donghwi Kim
Minkyoo Ji
Chi Soo Park
Jaeryong Lee
Dae Kyong Kim
Ha Hyung Kim
author_sort Jihye Kim
title Structural and Quantitative Characterization of Mucin-Type <i>O</i>-Glycans and the Identification of <i>O</i>-Glycosylation Sites in Bovine Submaxillary Mucin
title_short Structural and Quantitative Characterization of Mucin-Type <i>O</i>-Glycans and the Identification of <i>O</i>-Glycosylation Sites in Bovine Submaxillary Mucin
title_full Structural and Quantitative Characterization of Mucin-Type <i>O</i>-Glycans and the Identification of <i>O</i>-Glycosylation Sites in Bovine Submaxillary Mucin
title_fullStr Structural and Quantitative Characterization of Mucin-Type <i>O</i>-Glycans and the Identification of <i>O</i>-Glycosylation Sites in Bovine Submaxillary Mucin
title_full_unstemmed Structural and Quantitative Characterization of Mucin-Type <i>O</i>-Glycans and the Identification of <i>O</i>-Glycosylation Sites in Bovine Submaxillary Mucin
title_sort structural and quantitative characterization of mucin-type <i>o</i>-glycans and the identification of <i>o</i>-glycosylation sites in bovine submaxillary mucin
publisher MDPI AG
series Biomolecules
issn 2218-273X
publishDate 2020-04-01
description Bovine submaxillary mucin (BSM) is a gel-forming glycoprotein polymer, and Ser/Thr-linked glycans (<i>O</i>-glycans) are important in regulating BSM’s viscoelasticity and polymerization. However, details of <i>O</i>-glycosylation have not been reported. This study investigates the structural and quantitative characteristics of <i>O</i>-glycans and identifies <i>O</i>-glycosylation sites in BSM using liquid chromatography–tandem mass spectrometry. The <i>O</i>-glycans (consisting of di- to octa-saccharides) and their quantities (%) relative to total <i>O</i>-glycans (100%; 1.1 pmol per 1 μg of BSM) were identified with 14 major (>1.0%), 12 minor (0.1%–1.0%), and eight trace (<0.1%) <i>O</i>-glycans, which were characterized based on their constituents (sialylation (14 <i>O</i>-glycans; 81.9%, sum of relative quantities of each glycan), non-sialylation (20; 18.1%), fucosylation (20; 17.5%), and terminal-galactosylation (6; 3.6%)) and six core structure types [Gal-GalNAc, Gal-(GlcNAc)GalNAc, GlcNAc-GalNAc, GlcNAc-(GlcNAc)GalNAc, and GalNAc-GalNAc]. <i>O</i>-glycosylation sites were identified using <i>O</i>-glycopeptides (bold underlined; <sub>56</sub><b>S</b>GE<b>T</b>R<b>TS</b>VI, <sub>259</sub><b>S</b>H<b>SSS</b>GR<b>S</b>R<b>T</b>I, <sub>272</sub>G<b>S</b>P<b>SS</b>V<b>SS</b>AEQI, <sub>307</sub>RP<b>S</b>YGAL, <sub>625</sub>Q<b>T</b>LGPL, <sub>728</sub><b>T</b>M<b>TT</b>R<b>TS</b>VVV, and <sub>1080</sub>RPEDN<b>T</b>AVA) obtained from proteolytic BSM; these sites are in the four domains of BSM. The gel-forming mucins share common domain structures and glycosylation patterns; these results could provide useful information on mucin-type <i>O</i>-glycans. This is the first study to characterize <i>O</i>-glycans and identify <i>O</i>-glycosylation sites in BSM.
topic bovine submaxillary mucin
mucin-type O-glycan
O-glycosylation site
url https://www.mdpi.com/2218-273X/10/4/636
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