PURIFICATION AND PROPERTIES OF HUMAN INTESTINE ALANINE AMINOPEPTIDASE
PURIFICATION AND PROPERTIES OF HUMAN INTESTINE ALANINE AMINOPEPTIDASE
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DOI：
10.1016/0005-2744(80)90202-8
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发表时间：
1980-01-01
期刊：
影响因子：
--
通讯作者：
GARNER, CW
中科院分区：
文献类型：
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作者：
MCCLELLAN, JB;GARNER, CW
人肠丙氨酸氨基肽酶被纯化至大于90%的均一性。通过Triton X-100处理从粘膜细胞膜释放酶。天然酶在稀释缓冲液中的分子量为206，000，在十二烷基硫酸钠存在下的分子量为108，000。该酶被螯合剂抑制，表明酶中存在金属离子。测试的最有效的螯合剂抑制剂邻菲咯啉给出混合动力学（Ki = 67 μ M）。通过除去螯合剂恢复活性。该酶被具有疏水侧链的氨基酸如L-苯丙氨酸（Ki = 0.67 mM）竞争性抑制。嘌呤霉素和甲氧西林也抑制酶的竞争性（Ki = 12.5 mM）和非竞争性（Ki = 4.6 mM）的方式，分别。几种氨基酸β的动力学分析作为底物的萘酰胺显示出对具有疏水性或碱性氨基末端残基而不具有β-氨基的底物的偏好。分支L-甲硫氨酰-β-萘酰胺结合最紧密，而L-丙氨酰-β-萘酰胺水解最快。
Human intestinal alanine aminopeptidase was purified to greater than 90% homogeneity. The enzyme was released from mucosal cell membranes by Triton X-100 treatment. The native enzyme had a MW of 206,000 in dilute buffer and 108,000 in the presence of sodium dodecyl sulfate. The enzyme was inhibited by chelators suggesting the presence of a metal ion in the enzyme. The most potent chelator inhibitor tested, o-phenanthroline, gave mixed kinetics (Ki = 67 .mu.M). Activity was restored by removal of the chelator. The enzyme was inhibited competitively by amino acids having hydrophobic side chains such as L-phenylalanine (Ki = 0.67 mM). Puromycin and methicillin also inhibited the enzyme in the competitive (Ki = 12.5 mM) and noncompetitive (Ki = 4.6 mM) manner, respectively. Kinetic analysis of several amino acid .beta.-naphthylamides as substrates demonstrated the preference for substrates having hydrophobic or basic amino terminal residues with no .beta.-branching. L-Methionyl-.beta.-naphtyylamide was the most tightly bound while L-alanyl-.beta.-naphthylamide was the most rapidly hydrolyzed.