Purification and Characterization of Cysteine Conjugate Transaminases from Rat Liver
作者:Hiroki Tomisawa、Noriko Ichimoto、Yohko Takanohashi、Shigeyasu Ichihara、Hideo Fukazawa、Mitsuru Tateishi
DOI:10.3109/00498258809042224
日期:1988.1
1. Soluble cysteine-conjugate alpha-ketoglutarate transaminase (CAT-I) was purified about 670-fold from rat liver cytosol using s-(p-bromophenyl)-L-cysteine as amino acid substrate. The enzyme preparation of the final step of purification showed a single band in polyacrylamide gel electrophoresis. CAT-I accounted for 64% of the transaminase activity in cytosol. 2. The mol. wt of the enzyme was about
1.使用s-(对溴苯基)-L-半胱氨酸作为氨基酸底物,从大鼠肝细胞溶质中纯化可溶性半胱氨酸-缀合α-酮戊二酸转氨酶(CAT-1)约670倍。纯化的最后步骤的酶制剂在聚丙烯酰胺凝胶电泳中显示出一条条带。CAT-1占细胞溶胶中转氨酶活性的64%。2.摩尔。通过凝胶过滤测定,酶的wt%约为64,000。在Tris-乙酸盐缓冲液(pH 7.0)中,s-(对溴苯基)-L-半胱氨酸和α-酮戊二酸的Km值分别为1.0和1.3 mM。氨氧基乙酸,羟胺和KCN抑制了酶的活性。3.除CAT-1外,还从大鼠肝细胞溶质中部分纯化了两种同功酶(CAT-IIA和CAT-IIB)。关于摩尔。wt,对半胱氨酸缀合物的底物特异性,CAT-IIA和CAT-IIB与CAT-I非常相似。但是,在逆反应(半胱氨酸缀合物和α-酮戊二酸的形成反应)的速率和对L-天冬氨酸和L-半胱氨酸磺酸的底物特异性方面观察到这些酶的差异。