title: A strategy for fusion expression and preparation of functional glucagon-like peptide-1 (GLP-1) analogue by introducing an enterokinase cleavage site.
abstract: KGLP-1, a 31-amino acid glucagon-like peptide-1 (GLP-1) analogue, has a great therapeutic potential for anti-diabetes. In this work, a strategy for expression and purification of functional KGLP-1 peptide has been established. KGLP-1 cDNA was fused with glutathione S-transferase (GST), with an enterokinase cleavage site in the fusion junction. The recombinant fusion protein GST-KGLP-1 was affinity purified via the GST-tag, and then digested with enterokinase. The resulting GST part as well as the enzymes were eliminated by ultra-filtration followed by size exclusion chromatograph. The yield of purified KGLP-1 was approximately 12.1 mg/L, with purity of 96.18 %. The recombinant KGLP-1 was shown to have similar bioactivity as native GLP-1 when evaluated in a Chinese hamster ovary cell line expressing a GLP-1 receptor-egfp reporter gene.
authors.affiliation: College of Pharmacy and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, 94 Weijin Road, Tianjin, 300071, People's Republic of China, liuyang3513@163.com.
authors.forename: Yang
authors.initials: Y
authors.lastname: Liu
authors.forename: Limei
authors.initials: L
authors.lastname: Ren
authors.forename: Lingmiao
authors.initials: L
authors.lastname: Ge
authors.forename: Qingxin
authors.initials: Q
authors.lastname: Cui
authors.forename: Xiaofang
authors.initials: X
authors.lastname: Cao
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Hou
authors.forename: Fang
authors.initials: F
authors.lastname: Bai
authors.forename: Gang
authors.initials: G
authors.lastname: Bai
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001709:Biotechnology / Q000379:methods*; D016466:CHO Cells; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D004591:Electrophoresis, Polyacrylamide Gel; D004765:Enteropeptidase / Q000378:metabolism*; D052216:Glucagon-Like Peptide 1 / Q000031:analogs & derivatives* / Q000737:chemistry; D000067757:Glucagon-Like Peptide-1 Receptor; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D051381:Rats; D018027:Receptors, Glucagon / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: C000593835:Glp1r protein, rat; D000067757:Glucagon-Like Peptide-1 Receptor; D010455:Peptides; D018027:Receptors, Glucagon; D011993:Recombinant Fusion Proteins; D052216:Glucagon-Like Peptide 1; D004765:Enteropeptidase