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

内皮素（ET）是由21个氨基酸组成的强缩血管肽 (aa)，其最初是从培养的上清液中分离的， 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，优先影响 肾小管和血管。ET衍生自38 aa前体 肽，大ET（BET）。这种分裂是由一种身份不明的 蛋白酶、内皮素转换酶（ECE）。 我们试图确定：1）中性内肽酶EC. 3.4.24.11是否转化 BET至ET，使用重组中性内肽酶（rNEP），和2）如果 主动脉将BET转化为ET需要内皮 环. 125-I-ET-1与1 μ g/ml rNEP孵育， 肽在几分钟内，而其与10微克/毫升的 该酶在数秒内导致其完全裂解。两 phosphoramidon（10 μ M）和SQ-28，603（100 μ M）完全保护 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，样品中的大部分放射性计数作为完整洗脱 肽。与其对ET、phosphoramidon和SQ-28，603的作用相同 保护了rNEP对BET的降解。在研究期间的任何时间点， BET与rNEP的孵育在反应混合物中发现了ET。 在有内皮和无内皮的大鼠离体主动脉环中， BET-1引起剂量依赖性收缩。无显著 在存在或不存在的情况下，剂量-反应曲线的偏移 内皮细胞 这些结果表明：1）中性内肽酶EC. 3.4.24.11是一种 不是内皮素转换酶，2）内皮不需要 ET-1和BET-1的血管效应; 3）ECE存在于血管内皮细胞， 平滑肌

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.