Korean Journal of Agricultural Science (농업과학연구)

Institute of Agricultural Science, CNU (충남대학교 농업과학연구소)
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Recombinant production of human glucagon-like peptide-1 mutant

인간 Glucagon-like Peptide-1 변이체의 재조합 생산

  • Kim, Sung-Gun (Department of Biomedical Science, Youngdong University) ;
  • Park, Jong-Tae (Department of Food Science and Technology, Chungnam National University)
  • Received : 2014.05.30
  • Accepted : 2014.08.14
  • Published : 2014.09.30
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Abstract

Human Glucagon like peptide-1 (GLP-1) is an incretin hormone that promotes secretion of insulin. In order to eliminate the formation of the soluble aggregate, Ala19 in GLP-1 was substituted with Thr, resulting in a GLP-1 mutant GLP-1A19T. The gene synthesis of GLP-1A19T and the fusion of 6-lysine tagged ubiquitin gene were accomplished by using the overlap extension polymerase chain reaction. The ubiquitin fused GLP-1A19T (K6UbGLP-1A19T) is expressed as form of inclusion body with little formation of the soluble aggregation in recombinant E. coli. In order to produce K6UbGLP-1A19T in large amounts, fed-batch fermentation was carried out in a pH-stat feeding strategy. Maximum dry cell weight of 87.7 g/L and 20.4% of specific K6UbGLP-1A19T content were obtained. Solid-phase refolding using a cation exchanger was carried out to renature K6UbGLP-1A19T. The refolded K6UbGLP-1A19T aggregated little and was released GLP-1A19T by on-column cleavage with ubiquitin-specific protease-1. The molecular mass of GLP-1A19T showed an accurate agreement with its theoretical molecular mass.

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References

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