Production of a Bioactive Peptide (IIAEK) in Escherichia coli Using Soybean Proglycinin A1aB1b as a Carrier

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• 作者：Krisna Prak ; Shigeru Utsumi
• 刊名：Journal of Agricultural and Food Chemistry
• 出版年：2009
• 出版时间：May 13, 2009
• 年：2009
• 卷：57
• 期：9
• 页码：3792-3799
• 全文大小：987K
• 年卷期：v.57,no.9(May 13, 2009)
• ISSN：1520-5118

文摘

To produce large amounts of a peptide of fewer than 10 amino acid residues, construction of a gene encoding multimers of the small peptide is necessary. For this study a method was developed to facilitate the gene construction of high multimers of a small peptide with one step of cloning. A hypocholesterolemic peptide, IIAEK, from cow's milk β-lactoglobulin was used as a model peptide for the construction of a gene encoding multimers of IIAEK and for the production of the peptide. Two systems for direct expression of 28-mers of IIAEK sequences (28IIAEK) and expression of 34 IIAEK sequences (4 IIAEK sequences in each of the disordered regions I, II, and III and 14 and 8 IIAEK sequences in disordered regions IV and V, respectively) in a mutant of soybean proglycinin A1aB1b lacking 31 residues in disordered region IV [A1aB1b(Δ31)-34IIAEK] were used. The protein produced from both systems formed inclusion bodies. The expression level of A1aB1b(Δ31)-34IIAEK was 29.9% of the total cell proteins and that of the 28IIAEK was 2.0%. The insoluble A1aB1b(Δ31)-34IIAEK was digested by trypsin without any help from urea or chemicals, and the produced IIAEK was purified using an octadecyl silica column. The yield of IIAEK was 58.6%. The results showed that A1aB1b as a carrier of multiple peptides and use of an Escherichia coli expression system are suitable for production of bioactive peptide.