STUDIES ON A NEW PROTEOLYTIC-ENZYME FROM ACHROMOBACTER-LYTICUS M497-1 .1. PURIFICATION AND SOME ENZYMATIC-PROPERTIES
STUDIES ON A NEW PROTEOLYTIC-ENZYME FROM ACHROMOBACTER-LYTICUS M497-1 .1. PURIFICATION AND SOME ENZYMATIC-PROPERTIES
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DOI：
10.1016/0005-2744(81)90106-6
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发表时间：
1981-01-01
期刊：
影响因子：
--
通讯作者：
SOEJIMA, M
中科院分区：
文献类型：
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作者：
MASAKI, T;TANABE, M;SOEJIMA, M
A. lyticus M497-1在培养基中与溶菌酶一起产生3种碱性蛋白酶（蛋白酶I、蛋白酶II和蛋白酶III）。在这3种蛋白酶中，蛋白酶I （EC 3.4.21）。-)在分裂点的羧基侧显示赖氨酸残基的严格分裂。该酶经苯扎氯铵处理、丙酮分馏、cm -纤维素和deae -纤维素处理、AH-Sepharose 4B层析和等电聚焦法纯化。经聚丙烯酰胺凝胶电泳和超离心分析，该形态均相。酶的理化性质为MW 30,500；部分比容（.hivin）V), 0.717 ml/g；特性粘度（.eta.）， 0.0385 dl/g；等电点（pI） 6.9；和* *图形* *。在280 nm处，18.77。该酶每分子由294个氨基酸残基组成，其中甘氨酸为nh2末端，赖氨酸为cooh末端。酪蛋白、n -苯甲酰- dl -赖氨酸-对硝基苯胺和n -甲酰-l -赖氨酸甲酯的最佳pH值分别为8.5 ~ 10.7、9.0 ~ 9.5和7.8 ~ 8.2。DFP、苯甲磺酰氟和n - toyl -l -赖氨酸氯甲基酮对该酶有抑制作用，n - toyl -l -精氨酸氯甲基酮、EDTA -邻菲罗啉和PCMB对该酶无抑制作用。
A. lyticus M497-1 produces 3 kinds of alkaline proteases (protease I, II and III) in culture medium along with the bacteriolytic enzyme. Among these 3 proteases, protease I (EC 3.4.21.-) shows strict splitting for lysine residues at the carboxyl side of the splitting point. This enzyme was purified through a sequence of benzalkonium chloride treatment, acetone fractionation, CM-cellulose and DEAE-cellulose treatment, chromatography on AH-Sepharose 4B and isoelectric focusing method. This form was homogeneous by polyacrylamide gel electrophoresis and ultracentrifugation analysis. The physicochemical properties of the enzyme were MW 30,500; partial specific volume (.hivin.v), 0.717 ml/g; intrinsic viscosity (.eta.), 0.0385 dl/g; isoelectric point (pI) 6.9; and .**GRAPHIC**. at 280 nm, 18.77. The enzyme was composed of 294 residues of amino acid per molecule, with glycine as NH2-terminal and lysine as COOH-terminal amino acids. The optimum pH values with casein, N-benzoyl-DL-lysine-p-nitroanilide and N-tosyl-L-lysine methyl ester were 8.5-10.7, 9.0-9.5 and 7.8-8.2, respectively. The enzyme was inhibited by DFP, phenylmethanesulfonyl fluoride and N-tosyl-L-lysine chloromethyl ketone but not by N-tosyl-L-arginine chloromethyl ketone, EDTA o-phenanthroline and PCMB (p-chloromercuribenzoate).