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Tandem multimer expression of angiotensin I‐converting enzyme inhibitory peptide in Escherichia coli
Author(s) -
Fida Hasan M.,
Kumada Yoichi,
Terashima Masaaki,
Katsuda Tomohisa,
Katoh Shigeo
Publication year - 2009
Publication title -
biotechnology journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.144
H-Index - 84
eISSN - 1860-7314
pISSN - 1860-6768
DOI - 10.1002/biot.200800326
Subject(s) - recombinant dna , peptide , escherichia coli , biochemistry , chemistry , expression vector , microbiology and biotechnology , peptide sequence , inclusion bodies , gene , biology
It is common for small tandem peptide multimer genes to be indirectly inserted into expression vectors and fused with a protein tag. In this study, a multimer of the tandem angiotensin I‐converting enzyme inhibitory peptide (ACE‐IP) gene was directly transferred to a commercially available vector and the designed gene was expressed as a repeated peptide in Escherichia coli BL21(DE3)pLysS. The process further developed in our study was the construction of six‐repeated ACE‐IP synthetic genes and their direct insertion. Protein expression in inclusion bodies was confirmed by SDS‐PAGE and Western blot. Acid hydrolysis of inclusion bodies produced single‐unit peptides through cleavage of the aspartyl‐prolyl bonds. This cleaved recombinant peptide (rACE‐IP) was purified using immuno‐affinity chromatography followed by reversed phase‐HPLC. 105–115 mg of the lyophilized recombinant peptide was obtained from 1 L E. coli culture. In vitro biological activity of rACE‐IP was indistinguishable from that of the natural peptide produced by hydrolysis in artificial gastric juice or by acidic hydrolysis. The rACE‐IP prepared by recombinant DNA technology and solid‐phase synthesis methods showed a similar IC 50 . This strategy could be used for the expression of important peptides, which have N‐terminal proline (P) and C‐terminal aspartic acid residues (D) for commercial applications, e.g. functional foods and drinks.