A HIGHLY SENSITIVE FLUOROMETRIC ASSAY FOR ENKEPHALINASE, A NEUTRAL METALLOENDOPEPTIDASE THAT RELEASES TYROSINE-GLYCINE-GLYCINE FROM ENKEPHALINS
A HIGHLY SENSITIVE FLUOROMETRIC ASSAY FOR ENKEPHALINASE, A NEUTRAL METALLOENDOPEPTIDASE THAT RELEASES TYROSINE-GLYCINE-GLYCINE FROM ENKEPHALINS
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DOI：
10.1016/0003-2697(84)90425-1
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发表时间：
1984-01-01
影响因子：
2.9
通讯作者：
ROQUES, BP
中科院分区：
文献类型：
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作者：
FLORENTIN, D;SASSI, A;ROQUES, BP
合成了一种荧光肽Dansyl-D-Ala-Gly-Phe(PNO2)-Gly(DAGNPG)，作为脑啡肽代谢相关中性金属内肽酶(EC 3.4.24.11)的选择性底物。这种酶被命名为脑啡肽酶，它裂解DAGNPG的Gly-Phe-(PNO2)肽键(V[速度]=0.65微克/毫克蛋白质/分钟，Km=45微米)，导致荧光增强，这与硝基苯基对丹磺酰荧光的分子内猝灭有关。这一变化被用来定量测量不同组织中脑啡肽酶的活性，并测定各种化合物的抑制效力。底物不被氨基肽酶或二肽基层粘多肽酶活性切割，检测方法快速、方便、灵敏。
A fluorogenic peptide, dansyl-D-Ala-Gly-Phe(pNO2)-Gly (DAGNPG), was synthesized as a selective substrate for the neutral metalloendopeptidase (EC 3.4.24.11) involved in enkephalin metabolism. This enzyme, designated enkephalinase, cleaves the Gly-Phe-(pNO2) peptide bond of DAGNPG (V [velocity] = 0.65 .mu.mol/mg protein per min and Km= 45 .mu.M) leading to a fluorescence increase related to the disappearance of intramolecular quenching of the dansyl fluorescence by the nitrophenyl residue. This change was used for quantitative measurements of enkephalinase activity in different tissues and determination of inhibitory potency of various compounds. The substrate is not cleaved by aminopeptidase or dipeptidylaminopeptidase activities and the assay itself is rapid, convenient and sensitive.