Citation: ZHAO Yang, ZHANG Yong, WANG Mingchao, MENG Bo, YING Wantao, QIAN Xiaohong. Recombination of human galectin-3 with the carbohydrate recognition domain to achieve glycoprotein/glycopeptide enrichment[J]. Chinese Journal of Chromatography, ;2018, 36(12): 1197-1205. doi: 10.3724/SP.J.1123.2018.08017 shu

Recombination of human galectin-3 with the carbohydrate recognition domain to achieve glycoprotein/glycopeptide enrichment

ZHAO Yang ^1,2, ,
ZHANG Yong ² ,
WANG Mingchao ² ,
MENG Bo ² ,
YING Wantao ² ,
QIAN Xiaohong ^1,2

1.
College of Life Science and Bioengineering, Beijing University of Technology, Beijing, 100124, China;
2.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences(Beijing), Beijing Institute of Lifeomics, Beijing, 102206, China

Received Date: 14 August 2018

Fund Project: National Key R&D Program of China (Nos. 2017YFF0205400, 2016YFA0501300)

Though plant lectins have been widely used for the enrichment of glycoproteins/glycopeptides, mammalian lectins have rarely been explored for this purpose. Using the conserved sequence of the carbohydrate recognition domain (CRD) of the human galectin-3 protein, we engineered the following two novel human galectins:Gal3C (containing one CRD) and Tetra-Gal3C (containing four tandem CRDs). Moreover, we designed a galectin affinity column that could be used for glycoprotein/glycopeptide enrichment, by taking advantage of the ability of these recombinant galectins to become immobilized onto streptavidin agarose beads. SDS-PAGE, western blotting, and mass spectrometry were used to characterize the biological features and glycoprotein/glycopeptide enrichment abilities of both these recombinant galectins. Our results showed that both Gal3C and Tetra-Gal3C predominantly enrich glycoproteins/glycopeptides and that this enrichment process has a high specificity and sensitivity. Because Tetra-Gal3C contains three more CRD structural domains, it exhibited a better ability to achieve enrichment than Gal3C. Moreover, the immobilized Gal3C and Tetra-Gal3C were successfully used for the enrichment of glycoproteins from HepG2 cells, which indicated the selective glycoprotein/glycopeptide enrichment potential of recombinant galectins in complex samples.
- liquid chromatography-mass spectrometry (LC-MS),
- galectin-3 (Gal-3),
- carbohydrate recognition domain (CRD),
- glycoprotein enrichment
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