An integrated strategy for high-sensitive and multi-level glycoproteome analysis from low micrograms of protein samples

Gao, Weina1,2,3,4; Li, Hongjie1,2,3; Liu, Liping5; Huang, Peiwu2,3,6; Wang, Zhikun1,2,3; Chen, Wendong2,3; Ye, Mingliang4; Yu, Xiaofang5; Tian, Ruijun2,3

2019-08-30

10.1016/j.chroma.2019.04.041

JOURNAL OF CHROMATOGRAPHY A   影响因子和分区

0021-9673

1873-3778

Glycosylation, as a biologically important protein post-translational modification, often alters on both glycosites and glycans, simultaneously. However, most of current approaches focused on biased profiling of either glycosites or glycans, and limited by time-consuming process and milligrams of starting protein material. We describe here a simple and integrated spintip-based glycoproteomics technology (termed Glyco-SISPROT) for achieving a comprehensive view of glycoproteome with shorter sample processing time and low microgram starting material. By carefully integrating and optimizing SCX, C18 and Concanavalin A (Con A) packing material and their combination in spintip format, both predigested peptides and protein lysates could be processed by Glyco-SISPROT with high efficiency. More importantly, deglycopeptide, intact glycopeptide and glycans released by multiple glycosidases could be readily collected from the same Glyco-SISPROT workflow for LC-MS analysis. In total, above 1850 glycosites in (1) over tilde 770 unique deglycopeptides were characterized from mouse liver by using either 100 mu g of predigested peptides or directly using 100 mu g of protein lysates, in which about 30% of glycosites were released by both PNGase F and Endos. To the best of our knowledge, this approach should be one of the most comprehensive glycoproteomic approaches by using limited protein starting material. One significant benefit of Glyco-SISPROT is that whole processing time is dramatically reduced from a few days to less than 6 h with good reproducibility when protein lysates were directly processed by Glyco-SISPROT. We expect that this method will be suitable for multi-level glycoproteome analysis of rare biological samples with high sensitivity. (C) 2019 Elsevier B.V. All rights reserved.

Glycoproteomics Glycopeptide Glycan Integrated sample preparation LC-MS

SCI ; EI

英语

通讯

Guangdong Provincial Grants[2016A030312016] ; Guangdong Provincial Grants[2017B030301018]

Biochemistry & Molecular Biology ; Chemistry

Biochemical Research Methods ; Chemistry, Analytical

WOS:000472687800007

ELSEVIER SCIENCE BV

20191706834624

Peptides

Biology:461.9 ; Organic Compounds:804.1

CHEMISTRY

Web of Science

1.Harbin Inst Technol, Sch Chem & Chem Engn, Harbin 150080, Heilongjiang, Peoples R China
2.Southern Univ Sci & Technol, Dept Chem, Shenzhen 518055, Peoples R China
3.Southern Univ Sci & Technol, Guangdong Prov Key Lab Cell Microenvironm & Dis R, Shenzhen 518055, Peoples R China
4.Chinese Acad Sci, Dalian Inst Chem Phys, CAS Key Lab Separat Sci Analyt Chem, Dalian 116023, Peoples R China
5.Southern Univ Sci & Technol, Affiliated Hosp 1, Shenzhen Peoples Hosp, Shenzhen 518020, Peoples R China
6.Hong Kong Baptist Univ, Dept Chem, State Key Lab Environm & Biol Anal, Hong Kong, Peoples R China

Gao, Weina,Li, Hongjie,Liu, Liping,et al. An integrated strategy for high-sensitive and multi-level glycoproteome analysis from low micrograms of protein samples[J]. JOURNAL OF CHROMATOGRAPHY A,2019,1600:46-54.

Gao, Weina.,Li, Hongjie.,Liu, Liping.,Huang, Peiwu.,Wang, Zhikun.,...&Tian, Ruijun.(2019).An integrated strategy for high-sensitive and multi-level glycoproteome analysis from low micrograms of protein samples.JOURNAL OF CHROMATOGRAPHY A,1600,46-54.

Gao, Weina,et al."An integrated strategy for high-sensitive and multi-level glycoproteome analysis from low micrograms of protein samples".JOURNAL OF CHROMATOGRAPHY A 1600(2019):46-54.