Wagner G W, Tavianini M A, Herrmann K M, Dixon J E
Biochemistry. 1981 Jun 23;20(13):3884-90. doi: 10.1021/bi00516a034.
Rat brain enkephalin aminopeptidase was purified to apparent electrophoretic homogeneity. Enzyme activity was monitored during the purification by using ([3,5-3H2]Tyr)-Met-enkephalin and Tyr-beta-naphthylamide as substrates. It was shown that the enzyme activities resulting in hydrolysis of the tyrosine residue of ([3,5-3H2]Tyr)Met-enkephalin and formation of beta-naphthylamine from Tyr-beta-naphthylamide copurified. The homogeneous enzyme had a specific activity of 10.5 mumol of beta-naphthylamide hydrolyzed min-1 mg-1. Hydrolysis of Met-enkephalin yielded the products L-tyrosine and the tetrapeptide Gly-Gly-Phe-Met. Subsequent removal of glycine from Gly-Gly-Phe-Met was not observed with the purified enzyme. The homogeneous aminopeptidase has an apparent molecular weight of 115000 on Sephadex G-200 and a molecular weight of 102000 as determined by electrophoresis in the presence of sodium dodecyl sulfate. The enkephalin-degrading enzyme had a pH optimum of 6.5-7.0 and exhibited maximal activity at 40 degrees C. Enzyme activity was inhibited by metal chelators, and it was found that 1 mol of Zn2+ was associated with 1 mol of enzyme (102000 Mr). The enzyme hydrolyzes various neutral and basic amino acid beta-naphthylamides but will not utilize acidic, D-amino acid, or N-terminal-blocked amino acid beta-naphthylamides as substrates.
大鼠脑内啡肽氨基肽酶被纯化至表观电泳均一性。在纯化过程中，以([3,5 - 3H2]酪氨酸)-甲硫氨酸脑啡肽和酪氨酸-β-萘酰胺作为底物监测酶活性。结果表明，导致([3,5 - 3H2]酪氨酸)甲硫氨酸脑啡肽酪氨酸残基水解以及由酪氨酸-β-萘酰胺形成β-萘胺的酶活性共纯化。该均一酶的比活性为每分钟水解10.5 μmol β-萘酰胺每毫克。甲硫氨酸脑啡肽水解产生L-酪氨酸和四肽甘氨酰-甘氨酰-苯丙氨酰-甲硫氨酸。用纯化的酶未观察到甘氨酰-甘氨酰-苯丙氨酰-甲硫氨酸随后的甘氨酸去除。在Sephadex G - 200上，该均一氨基肽酶的表观分子量为115000，在十二烷基硫酸钠存在下通过电泳测定的分子量为102000。脑啡肽降解酶的最适pH为6.5 - 7.0，在40℃时表现出最大活性。酶活性受到金属螯合剂的抑制，并且发现1摩尔Zn2+与1摩尔酶（102000 Mr）相关。该酶水解各种中性和碱性氨基酸β-萘酰胺，但不将酸性、D-氨基酸或N-末端封闭的氨基酸β-萘酰胺作为底物。
