Isolation and characterization of DL-methylmalonyl-coenzyme A racemase from rat liver.
Isolation and characterization of DL-methylmalonyl-coenzyme A racemase from rat liver.
复制标题
大鼠肝脏 DL-甲基丙二酰辅酶 A 消旋酶的分离和表征。
DOI：
10.1016/0003-9861(85)90381-9
复制
发表时间：
1985
影响因子：
3.9
通讯作者：
Allen,RH
中科院分区：
文献类型：
--
作者：
Stabler,SP;Marcell,PD;Allen,RH
某些氨基酸和其它化合物通过丙酰-辅酶A-甲基丙二酰-辅酶A-甲基丙二酰-辅酶A琥珀酰-辅酶A参与三羧酸循环代谢。d-甲基丙二酰-辅酶A也可以通过不作用于l-甲基丙二酰-辅酶A的特异性水解酶转化为甲基丙二酸和辅酶A [R. J. Kovachy，S. D. Copley和R. H.艾伦（1983）生物化学杂志258，11415 -11421]。由于对大鼠肝消旋酶知之甚少，并且由于它参与了d-甲基丙二酰-CoA →甲基丙二酰-CoA →三羧酸循环（相对于甲基丙二酸）的流动，我们开发了一种新的测定方法，并将大鼠肝消旋酶纯化23，000倍至均一。消旋酶的分子量为32，000，它含有两个Mr 16，000的亚基，它们不是通过二硫键连接的。大鼠肝脏与大鼠和人白色血细胞消旋酶具有免疫相关性。EDTA可使其完全失活，加入Co+2可使其活化，浓度为0.2 μm时活化率为50%。最大活化的较低水平得到较高浓度的Co+3，Fe+2，和Mn+2。其它金属离子如Zn+2、Cu+2、Cu+1和Cd+2在等浓度的Co+2存在下也能完全抑制消旋酶。纯化的消旋酶似乎结合1 mol Co/mol亚基。
Certain amino acids and other compounds are metabolized via propionyl-CoAd-methylmalonyl-CoAl-methylmalonyl-CoAsuccinyl-CoA ↔ tricarboxylic acid cycle.d-Methylmalonyl-CoA can also be converted to methylmalonic acid and coenzyme A by a specific hydrolase that does not act onl-methylmalonyl-CoA [R. J. Kovachy, S. D. Copley, and R. H. Allen (1983)J. Biol. Chem.258,11415–11421]. Because little is known about mammaliandl-methylmalonyl-CoA racemase and because it is involved in the flow ofd-methylmalonyl-CoA tol-methylmalonyl-CoA → tricarboxylic acid cycle (versus to methylmalonic acid), we developed a new assay and purified rat liver racemase 23,000-fold to homogeneity. The molecular weight of the racemase is 32,000 and it contains two subunits ofMr16,000 that are not connected by disulfide bonds. The rat liver and the rat and human white blood cell racemase are immunologically related. They are completely inactivated by EDTA and can be activated by the addition of Co+2, with 50% activation occurring at a concentration of 0.2 μm. Lower levels for maximal activation were obtained with higher concentrations of Co+3, Fe+2, and Mn+2. Other metals such as Zn+2, Cu+2, Cu+1, and Cd+2completely inhibited racemase even in the presence of equal concentrations of Co+2. The purified racemase appears to bind 1 mol Co/mol subunit.