Endoglycosidase-based Chemoenzymatic Synthesis of Homogeneous Glycopeptides and Glycoproteins Carrying Defined N-Glycans

China Biotechnology

2012, Vol. 32

Issue (02): 107-116 DOI:

Endoglycosidase-based Chemoenzymatic Synthesis of Homogeneous Glycopeptides and Glycoproteins Carrying Defined N-Glycans

ZOU Yang^1,2, XUE Meng-yang^1,2, ZHAO Ying-hua¹, CHEN Min^1,2

1. The State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China;
2. National Glycoengineering Research Center, Shandong University, Jinan 250100, China

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Abstract

Protein glycosylation is the important step involved in post-translational modifications within eukaryotes, and it plays a significant role in protein folding and transportational processes. The oligosaccharide moieties of glycoconjugates have a major impact on the structure and functions of protein, and influence biological phenomena such as lectin binding, viral infection and cellular recognition. In the field of glycobiology, it is necessary to investigate the homogeneous glycopeptides and glycoprotein for detailed structural and functional studies of glycoproteins. As a result, how to synthesize this kind of glycoprotein and glycopeptides efficiently has become an important subject in glycotechnology. At the present time, three major strategies have been developed to synthesize homogeneous glycopeptides. One is using pre-formed glycosyl amino acids as building blocks in solid-phase or solution-phase peptide synthesis. Another strategy is combining an oligosaccharide glycosylamine with a pre-assembled polypeptide which contains a free or selectively activated aspartyl side chain to synthesize glycoprotein. The main disadvantages of these approaches are that the strong acidic conditions may cause partial hydrolysis of the attached oligosaccharide moiety. To deal with these problems mentioned in the above strategies, a new chemoenzymatic approach combined with endoglycosidase which from microorganisms to synthesize homogeneous glycopeptides is developed. This approach requires only monosaccharide-tagged polypeptides, and the reaction condition is very moderate. The sugar chain extension is combined with free polypeptides in aqueous solutions, without the using of protecting groups. Both glycosyltransferases and endoglycosidases have been explored for this chemoenzymatic strategy. Compared with glycosyltransferase-based methods, the endoglycosidase-based approach is more attractive because it can connect a large intact oligosaccharide to a GlcNAc polypeptide in one-step. Although this endoglycosidase-based chemoenzymatic approach is better than other methods, it still has shortcomings, such as low transglycosylation efficiency and product hydrolysis issue. The synthesis of these homogeneous glycopeptides is still a challenging work because of every strategy has its own advantages and limitations. But compared with other strategies, endoglycosidase-based chemoenzymatic synthetic approach is more attractive owing to its faster developing and more practical significance. Following aspects are focused on: the research progress of endoglycosidase-based chemoenzymatic synthetic strategy, including 4 aspects: (1) Studies on the catalytic mechanism of endoglycosidase. Such as substrate-assisted mechanism. (2) Researches of glycosyl donors of endoglycosidase. Based on the substrate-assisted mechanism of ENGase-catalyzed reaction proceed, sugar oxazolines was used as activated substrates for ENGase-catalyzed transglycosylation, and this substrate surprisingly induced substantial improvement of the synthetic efficiency and high-yield of large glycopeptides. (3) Investigations of mutants of endoglycosidase. Which lack hydrolytic activity but can still combine an activated glycosyl donor to peptide acceptor for catalysis to form a new glycosidic bond. (4) The applications of glycoprotein and glycopeptides which have homogeneous sugar chains and synthesized by endoglycosidase-based chemoenzymatic approaches. These glycoproteins could be used in the study of structure-function relationship of protein, and the study of modifications of protein glycosylation.

Key words： Glycoprotein Glycopeptides Chemoenzymatic synthesis Endo-β-N-acetylglucosaminidase Oligosaccharide oxazoline

Received: 09 November 2011 Published: 25 February 2012

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Cite this article:

ZOU Yang, XUE Meng-yang, ZHAO Ying-hua, CHEN Min. Endoglycosidase-based Chemoenzymatic Synthesis of Homogeneous Glycopeptides and Glycoproteins Carrying Defined N-Glycans. China Biotechnology, 2012, 32(02): 107-116.

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https://manu60.magtech.com.cn/biotech/ OR https://manu60.magtech.com.cn/biotech/Y2012/V32/I02/107

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