Nakagawa Tsutomu, Akaki Jyunji, Satou Ryousuke, Takaya Masatoshi, Iwata Hideyuki, Katsurada Akemi, Nishiuchi Kazuhiro, Ohmura Yoshihiro, Suzuki Fumiaki, Nakamura Yukio
Laboratory of Applied Biochemistry, Faculty of Applied Biological Science, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.
Biol Chem. 2007 Feb;388(2):237-46. doi: 10.1515/BC.2007.026.
The amino acid sequence His-Pro-Phe as N-terminal residues 6-8 of the natural renin substrate, angiotensinogen, is conserved among species. We investigated whether this His-Pro-Phe motif functions as the determinant of the substrate specificity of renin. Mutant angiotensinogens in which the Ile-His-Pro-Phe-His-Leu sequence at positions 5-10 of wild-type angiotensinogen was replaced by either His-Pro-Phe-His-Leu-Leu or Ala-Ile-His-Pro-Phe-His were cleaved by renin at the C-terminal side of residues 9 and 11, respectively, while wild-type angiotensinogen was cleaved at residue 10. A triple Ala substitution for the His-Pro-Phe motif of angiotensinogen prevented its cleavage by renin. In contrast, triple Ala substitution for residues 9-11, including the natural site of cleavage by renin, allowed cleavage between the two Ala residues at positions 10 and 11. Furthermore, the 33-residue C-terminal peptide of human megsin, which carries a naturally occurring His-Pro-Phe sequence, was cleaved by renin at the C-terminal side of the His-Pro-Phe-Leu-Phe sequence. These results indicate that the His-Pro-Phe motif of angiotensinogen is a crucial determinant of the substrate specificity of renin. By binding to a corresponding pocket on renin, the His-Pro-Phe motif may act as a molecular anchor to recruit the scissile peptide bond to a favorable site for catalysis.
天然肾素底物血管紧张素原的 N 端第 6 - 8 位氨基酸序列 His - Pro - Phe 在物种间保守。我们研究了该 His - Pro - Phe 基序是否作为肾素底物特异性的决定因素。将野生型血管紧张素原第 5 - 10 位的 Ile - His - Pro - Phe - His - Leu 序列分别替换为 His - Pro - Phe - His - Leu - Leu 或 Ala - Ile - His - Pro - Phe - His 的突变型血管紧张素原，分别在第 9 位和第 11 位残基的 C 端被肾素切割，而野生型血管紧张素原在第 10 位残基处被切割。血管紧张素原的 His - Pro - Phe 基序的三个 Ala 取代阻止了其被肾素切割。相反，对包括肾素天然切割位点在内的第 9 - 11 位残基进行三个 Ala 取代，允许在第 10 位和第 11 位的两个 Ala 残基之间进行切割。此外，携带天然 His - Pro - Phe 序列的人 megsin 的 33 个残基的 C 端肽在 His - Pro - Phe - Leu - Phe 序列的 C 端被肾素切割。这些结果表明血管紧张素原的 His - Pro - Phe 基序是肾素底物特异性的关键决定因素。通过与肾素上的相应口袋结合，His - Pro - Phe 基序可能作为分子锚，将可裂解肽键募集到有利于催化的位点。
