Optimal designing of beta-conglycinin to genetically incorporate RPLKPW, a potent anti-hypertensive peptide.
Previously, we introduced the RPLKPW sequence, a highly potent hypotensive peptide designed based on ovokinin (2-7), into three homologous sites in the soybean beta-conglycinin alpha' subunit by site-directed mutagenesis. The modified protein expressed in Escherichia coli reduced blood pressure of spontaneously hypertensive rats (SHRs) after oral administration at a dose of 10 mg/kg, which suggested about 30% of the introduced peptide was released in vivo. In this study amino acid residues around the RPLKPW sequence were optimized with a use of synthetic peptides to facilitate release of RPLKPW by gastrointestinal proteases. Then, fourth RPLKPW was also introduced into the extension domain of the protein. The newly modified protein, which was produced in E. coli, significantly lowered blood pressure in SHRs at a dose of 2.5 mg/kg 4 h after oral administration. Furthermore, we produced an extension domain that corresponds to residues 1-143 of the modified alpha' subunit containing four RPLKPW sequences by introducing a termination codon. The minimum effective dose of the modified extension domain was 1.0 mg/kg, which is 1/2000 that of ovalbumin.
Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Gokasho Uji, Kyoto 611-0011, Japan.
Kunihiko
K
Onishi
Nobuyuki
N
Matoba
Yuko
Y
Yamada
Naomi
N
Doyama
Nobuyuki
N
Maruyama
Shigeru
S
Utsumi
Masaaki
M
Yoshikawa
D000284:Administration, Oral; D000818:Animals; D052179:Antigens, Plant; D000959:Antihypertensive Agents / Q000008:administration & dosage / Q000737:chemistry / Q000494:pharmacology; D002852:Chromatography, Ion Exchange; D015195:Drug Design; D005916:Globulins / Q000008:administration & dosage / Q000737:chemistry* / Q000494:pharmacology; D008297:Male; D016297:Mutagenesis, Site-Directed; D010047:Ovalbumin / Q000737:chemistry; D051381:Rats; D011918:Rats, Inbred SHR; D055314:Seed Storage Proteins; D030262:Soybean Proteins / Q000008:administration & dosage / Q000737:chemistry* / Q000494:pharmacology; D013997:Time Factors
D052179:Antigens, Plant; D000959:Antihypertensive Agents; D005916:Globulins; D055314:Seed Storage Proteins; D030262:Soybean Proteins; C013894:beta-conglycinin protein, Glycine max; D010047:Ovalbumin