Expression and Characterization of Human β-1, 4-Galactosyltransferase 1 (β4GalT1) Using Silkworm–Baculovirus Expression System

Handle URI:
http://hdl.handle.net/10754/623835
Title:
Expression and Characterization of Human β-1, 4-Galactosyltransferase 1 (β4GalT1) Using Silkworm–Baculovirus Expression System
Authors:
Morokuma, Daisuke; Xu, Jian; Hino, Masato; Mon, Hiroaki; Merzaban, Jasmeen S. ( 0000-0002-7276-2907 ) ; Takahashi, Masateru; Kusakabe, Takahiro; Lee, Jae Man
Abstract:
Baculovirus expression vector system (BEVS) is widely known as a mass-production tool to produce functional recombinant glycoproteins except that it may not be always suitable for medical practice due to the differences in the structure of N-linked glycans between insects and mammalian. Currently, various approaches have been reported to alter N-linked glycan structures of glycoproteins derived from insects into terminally sialylated complex-type N-glycans. In the light of those studies, we also proposed in vitro maturation of N-glycan with mass-produced and purified glycosyltransferases by silkworm–BEVS. β-1,4-Galactosyltransferase 1 (β4GalT1) is known as one of type II transmembrane enzymes that transfer galactose in a β-1, 4 linkage to accepter sugars, and a key enzyme for further sialylation of N-glycans. In this study, we developed a large-scale production of recombinant human β4GalT1 (rhβ4GalT1) with N- or C-terminal tags in silkworm–BEVS. We demonstrated that rhβ4GalT1 is N-glycosylated and without mucin-type glycosylation. Interestingly, we found that purified rhβ4GalT1 from silkworm serum presented higher galactosyltransferase activity than that expressed from cultured mammalian cells. We also validated the UDP-galactose transferase activity of produced rhβ4GalT1 proteins by using protein subtracts from silkworm silk gland. Taken together, rhβ4GalT1 from silkworms can become a valuable tool for producing high-quality recombinant glycoproteins with mammalian-like N-glycans.
KAUST Department:
Biological and Environmental Sciences and Engineering (BESE) Division; Laboratory of DNA Replication and Recombination
Citation:
Morokuma D, Xu J, Hino M, Mon H, Merzaban JS, et al. (2017) Expression and Characterization of Human β-1, 4-Galactosyltransferase 1 (β4GalT1) Using Silkworm–Baculovirus Expression System. Molecular Biotechnology 59: 151–158. Available: http://dx.doi.org/10.1007/s12033-017-0003-1.
Publisher:
Springer Nature
Journal:
Molecular Biotechnology
Issue Date:
24-Mar-2017
DOI:
10.1007/s12033-017-0003-1
Type:
Article
ISSN:
1073-6085; 1559-0305
Additional Links:
http://link.springer.com/article/10.1007/s12033-017-0003-1
Appears in Collections:
Articles; Biological and Environmental Sciences and Engineering (BESE) Division

Full metadata record

DC FieldValue Language
dc.contributor.authorMorokuma, Daisukeen
dc.contributor.authorXu, Jianen
dc.contributor.authorHino, Masatoen
dc.contributor.authorMon, Hiroakien
dc.contributor.authorMerzaban, Jasmeen S.en
dc.contributor.authorTakahashi, Masateruen
dc.contributor.authorKusakabe, Takahiroen
dc.contributor.authorLee, Jae Manen
dc.date.accessioned2017-05-31T11:23:08Z-
dc.date.available2017-05-31T11:23:08Z-
dc.date.issued2017-03-24en
dc.identifier.citationMorokuma D, Xu J, Hino M, Mon H, Merzaban JS, et al. (2017) Expression and Characterization of Human β-1, 4-Galactosyltransferase 1 (β4GalT1) Using Silkworm–Baculovirus Expression System. Molecular Biotechnology 59: 151–158. Available: http://dx.doi.org/10.1007/s12033-017-0003-1.en
dc.identifier.issn1073-6085en
dc.identifier.issn1559-0305en
dc.identifier.doi10.1007/s12033-017-0003-1en
dc.identifier.urihttp://hdl.handle.net/10754/623835-
dc.description.abstractBaculovirus expression vector system (BEVS) is widely known as a mass-production tool to produce functional recombinant glycoproteins except that it may not be always suitable for medical practice due to the differences in the structure of N-linked glycans between insects and mammalian. Currently, various approaches have been reported to alter N-linked glycan structures of glycoproteins derived from insects into terminally sialylated complex-type N-glycans. In the light of those studies, we also proposed in vitro maturation of N-glycan with mass-produced and purified glycosyltransferases by silkworm–BEVS. β-1,4-Galactosyltransferase 1 (β4GalT1) is known as one of type II transmembrane enzymes that transfer galactose in a β-1, 4 linkage to accepter sugars, and a key enzyme for further sialylation of N-glycans. In this study, we developed a large-scale production of recombinant human β4GalT1 (rhβ4GalT1) with N- or C-terminal tags in silkworm–BEVS. We demonstrated that rhβ4GalT1 is N-glycosylated and without mucin-type glycosylation. Interestingly, we found that purified rhβ4GalT1 from silkworm serum presented higher galactosyltransferase activity than that expressed from cultured mammalian cells. We also validated the UDP-galactose transferase activity of produced rhβ4GalT1 proteins by using protein subtracts from silkworm silk gland. Taken together, rhβ4GalT1 from silkworms can become a valuable tool for producing high-quality recombinant glycoproteins with mammalian-like N-glycans.en
dc.publisherSpringer Natureen
dc.relation.urlhttp://link.springer.com/article/10.1007/s12033-017-0003-1en
dc.subjectBaculovirus expression vector systemen
dc.subjectβ-1, 4-Galactosyltransferase 1en
dc.subjectN-glycosylation Silkwormen
dc.titleExpression and Characterization of Human β-1, 4-Galactosyltransferase 1 (β4GalT1) Using Silkworm–Baculovirus Expression Systemen
dc.typeArticleen
dc.contributor.departmentBiological and Environmental Sciences and Engineering (BESE) Divisionen
dc.contributor.departmentLaboratory of DNA Replication and Recombinationen
dc.identifier.journalMolecular Biotechnologyen
dc.contributor.institutionLaboratory of Insect Genome Science, Kyushu University Graduate School of Bioresource and Bioenvironmental Sciences, Fukuoka, Japanen
dc.contributor.institutionLaboratory of Insect Genome Science, Kyushu University Graduate School of Bioresource and Bioenvironmental Sciences, Fukuoka, Japan|en
kaust.authorMerzaban, Jasmeen S.en
kaust.authorTakahashi, Masateruen
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