Nurun Nabi A H M, Uddin Nasir M, Nakagawa Tsutomu, Iwata Hideyuki, Ichihara Atsuhiro, Inagami Tadashi, Suzuki Fumiaki
Graduate School of Agricultural Science, Gifu University, Gifu, Japan.
Front Biosci. 2007 Sep 1;12:4810-7. doi: 10.2741/2429.
A role of the "handle" region in the prorenin prosegment sequence was investigated to demonstrate the crucial non-proteolytic activation of prorenin by binding to the recombinant (pro)renin receptor on the COS-7 cell membrane. The plasmid DNA containing either rat or human (pro)renin receptor was transfected into the COS-7 cells. The highest amount of receptor was observed on the COS-7 cell membrane after 18 h transfection. Of the total rat and human prorenin, 90% and 50% were bound to each of the respective receptors, respectively. The Kd values were 0.89 and 1.8 nM, respectively. Rat prorenin was activated non-proteolytically by the receptor. The Km was determined 1.0 microM when sheep angiotensinogen was used as the substrate. Human prorenin was also activated by the receptor. The Km was 0.71 microM. Additionally, decapeptides (10P-19P) known as "decoy" peptide and pentapeptides (11P-15P) named "handle" region peptide, were observed to inhibit the binding of both prorenins to receptors, respectively. The Ki were similar around 7 nM for both the peptides. Other two region peptides in the prosegment did not interfere the binding. These results show that the "handle" region probably plays a crucial role in prorenin binding to the receptor and in its enzymic activity by non-proteolytic activation.
研究了肾素原前体序列中“柄”区的作用，以证明肾素原通过与COS-7细胞膜上的重组（前）肾素受体结合而进行的关键非蛋白水解激活。将含有大鼠或人（前）肾素受体的质粒DNA转染到COS-7细胞中。转染18小时后，在COS-7细胞膜上观察到最高量的受体。在总的大鼠和人肾素原中，分别有90%和50%与各自的受体结合。Kd值分别为0.89和1.8 nM。大鼠肾素原被受体非蛋白水解激活。当使用绵羊血管紧张素原作为底物时，测定的Km为1.0 microM。人肾素原也被受体激活。Km为0.71 microM。此外，观察到被称为“诱饵”肽的十肽（10P-19P）和被称为“柄”区肽的五肽（11P-15P）分别抑制两种肾素原与受体的结合。两种肽的Ki相似，约为7 nM。前体中的其他两个区域肽不干扰结合。这些结果表明，“柄”区可能在肾素原与受体结合及其通过非蛋白水解激活的酶活性中起关键作用。
