METABOLISM OF ENDOTHELIN AND BIG ENDOTHELIN--INVOLVEMENT OF NEUTRAL ENDOPEPTIDASE

内皮素(ET)是一种由21个氨基酸组成的强效血管收缩多肽。 (Aa)，最初从培养上清液中分离出来 内皮细胞。除了已知的血管收缩作用外，ET 发挥广泛的生物作用，对 肾小管和血管。ET来源于38个氨基酸的前体 肽，大ET(BET)。这个卵裂是由一个身份不明的人执行的 蛋白水解酶、内皮素转换酶(ECEs)。 我们试图确定：1)中性内肽酶EC.3.4.24.11是否会转化为 最好是ET，使用重组中性内切酶(RNEP)，以及2)如果 主动脉将BET转化为ET需要内皮细胞 戒指。 125-I-ET-1与1微克/毫升rNEP孵育可引起细胞的降解 几分钟内，而它与10微克/毫升的 这种酶在几秒钟内就完全裂解了。两者都有 磷酰胺(10微米)和SQ-28,603(100微米)完全受保护 ET-1被rNEP降解。~(125)I-BET-1的降解率较高 比ET-1慢。即使在与10微克/毫升的 酶，样本中的大部分放射性计数洗脱后完好无损。 多肽。对ET、磷酰胺和SQ-28,603的影响相同 保护了rNEP对BET的降解。在此期间的任何时间点 在反应混合物中发现了BET与rNEP的孵育。 在有内皮和无内皮的大鼠离体主动脉环中，ET-1和ET-1 BET-1引起剂量依赖性收缩。没有显著的 在存在或不存在的情况下剂量-反应曲线的移动 内皮细胞。 这些发现表明：1)中性内肽酶EC.3.4.24.11是 不是内皮素转换酶，2)内皮不是必需的 ET-1和BET-1的血管效应；3)ECEs存在于血管内 平滑的肌肉。

Endothelin (ET) is a potent vasoconstrictor peptide of 21 amino- acids (aa), which was originally isolated from the supernatant of cultured endothelial cells. In addition to its known vasoconstricting effects, ET exerts a wide range of biological actions, with preferential effects on renal tubules and blood vessels. ET is derived from a 38 aa precursor peptide, Big ET (BET). This cleavage is performed by an unidentified protease, endothelin converting enzyme (ECE). We sought to determine: 1) if neutral endopeptidase EC.3.4.24.11 converts BET to ET, using recombinant neutral endopeptidase (rNEP), and 2) if endothelium is required for the conversion of BET to ET, by aortic rings. Incubation of 125-I-ET-1 with 1microg/ml rNEP resulted in degradation of the peptide within minutes, whereas its incubation with 10 microg/ml of the enzyme resulted in its total cleavage within seconds. Both phosphoramidon (10 microM) and SQ-28,603 (100microM) fully protected ET-1 from degradation by rNEP. The degradation of 125-I-BET-1 was much slower than that of ET-1. Even after incubation with 10 microg/ml of the enzyme, most of radioactive counts in the sample eluted as intact peptide. Identical to their effect on ET, phosphoramidon and SQ-28,603 protected the degradation of BET by rNEP. At no time point during the incubation of BET with rNEP, was ET found in the reaction mixture. In isolated rat aortic rings with and without the endothelium, both ET-1 and BET-1 caused dose- dependent contractions. There was no significant shift of the dose-response curve in the presence or absence of the endothelium. These findings indicate that: 1) neutral endopeptidase EC.3.4.24.11 is not an endothelin converting enzyme, 2) endothelium is not required for the vascular effects of ET-1 and BET-1, and 3) ECE exists in vascular smooth muscle.