人肾组织组织蛋白酶H的一种新纯化方法、其性质及动力学数据。

A new purification procedure of human kidney cathepsin H, its properties and kinetic data.

作者信息

Popović T, Brzin J, Kos J, Lenarcic B, Machleidt W, Ritonja A, Hanada K, Turk V

机构信息

Odsek za biokemijo, Institut J. Stefan, Univerza E. Kardelja, Ljubljana, Jugoslavia.

出版信息

Biol Chem Hoppe Seyler. 1988 May;369 Suppl:175-83.

PMID:3202963

Abstract

A purification procedure of cathepsin H from human kidney is presented. It includes gel filtration, ion exchange chromatography, and covalent chromatography on thiol Sepharose as an essential step. Purified cathepsin H emerges in an isoelectric focusing gel at pH 6.1 and 6.3. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate shows a molecular mass of about 28 kDa. Less than 20% of the enzyme preparation can be separated into a heavy (24 kDa) and a light chain (4 kDa) after reduction and gel filtration on Sephacryl S-200. The partial amino-acid sequence of human cathepsin H shows its close similarity to rat cathepsin H. Inhibition constants (Ki) of cathepsins H and B with chicken cystatin, two forms of human stefin A, human stefin B, and two forms of human cystatin C are in the range of 10(-9) to 10(-11)M.

摘要

本文介绍了一种从人肾中纯化组织蛋白酶H的方法。该方法包括凝胶过滤、离子交换色谱和以硫醇琼脂糖凝胶为关键步骤的共价色谱。纯化后的组织蛋白酶H在等电聚焦凝胶中pH值为6.1和6.3处出现。在十二烷基硫酸钠存在下的聚丙烯酰胺凝胶电泳显示其分子量约为28 kDa。还原后在Sephacryl S - 200上进行凝胶过滤，不到20%的酶制剂可分离为重链（24 kDa）和轻链（4 kDa）。人组织蛋白酶H的部分氨基酸序列显示其与大鼠组织蛋白酶H高度相似。组织蛋白酶H和B对鸡半胱氨酸蛋白酶抑制剂、两种形式的人丝抑素A、人丝抑素B以及两种形式的人半胱氨酸蛋白酶抑制剂C的抑制常数（Ki）在10^(-9)至10^(-11)M范围内。