Improvement of Endo-β-N-acetylglucosaminidase H production using silkworm-baculovirus protein expression system

Atsushi Masuda, Jian Xu, Takumi Mitsudome, Daisuke Morokuma, Hiroaki Mon, Yutaka Banno, Takahiro Kusakabe, Man Lee

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Endo-β-. N-acetylglucosaminidase H (Endo H) catalyzes cleavage between the GlcNAc residues of the chitobiose core of N-linked glycans, leaving one GlcNAc residues attached to asparagine. Endo H cleaves high mannose and hybrid, but not complex, N-linked oligosaccharides on glycoproteins. Because of its unique specificity, Endo H is widely used for the structural and functional analyses of glycoproteins. In our previous study, the recombinant Endo H was produced as a secreted protein using silkworm-baculovirus expression system, but the yield was low (30. μg Endo H/10. ml larval hemolymph) compared to that of Escherichia coli. In this study, we purified active recombinant Endo H as an intracellular protein from fat body of silkworm infected with the recombinant baculovirus expressing Endo H without the exogenous signal peptide. Remarkably, the yield (9.3. mg from 20 silkworm larvae) was about 310-fold higher than that secreted into larval hemolymph as reported previously. In addition, we screened the silkworm strains maintained in Kyushu University and identified n17 as a high-level expression strain for Endo H.

Original languageEnglish
Pages (from-to)175-180
Number of pages6
JournalJournal of Asia-Pacific Entomology
Volume18
Issue number2
DOIs
Publication statusPublished - Jan 1 2015

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Baculoviridae
silkworms
protein synthesis
hemolymph
glycoproteins
fat body
asparagine
signal peptide
mannose
oligosaccharides
insect larvae
polysaccharides
proteins
Japan
Escherichia coli

All Science Journal Classification (ASJC) codes

  • Insect Science

Cite this

Improvement of Endo-β-N-acetylglucosaminidase H production using silkworm-baculovirus protein expression system. / Masuda, Atsushi; Xu, Jian; Mitsudome, Takumi; Morokuma, Daisuke; Mon, Hiroaki; Banno, Yutaka; Kusakabe, Takahiro; Lee, Man.

In: Journal of Asia-Pacific Entomology, Vol. 18, No. 2, 01.01.2015, p. 175-180.

Research output: Contribution to journalArticle

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