Production of a bioactive peptide (IIAEK) in Escherichia coli using soybean proglycinin A1ab1b as a carrier.

Krisna Prak, Shigeru Utsumi

Journal of agricultural and food chemistry 2009 May 13

filter terms:

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 beta-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(Delta31)-34IIAEK] were used. The protein produced from both systems formed inclusion bodies. The expression level of A1aB1b(Delta31)-34IIAEK was 29.9% of the total cell proteins and that of the 28IIAEK was 2.0%. The insoluble A1aB1b(Delta31)-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.

Citation

Krisna Prak, Shigeru Utsumi. Production of a bioactive peptide (IIAEK) in Escherichia coli using soybean proglycinin A1ab1b as a carrier. Journal of agricultural and food chemistry. 2009 May 13;57(9):3792-9