title: Tandem multimer expression of angiotensin I-converting enzyme inhibitory peptide in Escherichia coli.
abstract: 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.
authors.affiliation: Department of Molecular Science and Material Engineering, Graduate School of Science and Technology, Faculty of Engineering, Kobe University, Kobe, Japan.
authors.forename: Hasan M
authors.initials: HM
authors.lastname: Fida
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Kumada
authors.forename: Masaaki
authors.initials: M
authors.lastname: Terashima
authors.forename: Tomohisa
authors.initials: T
authors.lastname: Katsuda
authors.forename: Shigeo
authors.initials: S
authors.lastname: Katoh
mesh_terms: D000806:Angiotensin-Converting Enzyme Inhibitors / Q000378:metabolism*; D019281:Dimerization; D004926:Escherichia coli / Q000502:physiology*; D010455:Peptides / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000378:metabolism*
chemical_list: D000806:Angiotensin-Converting Enzyme Inhibitors; D010455:Peptides; D011994:Recombinant Proteins