Chan W W
J Biol Chem. 1975 Jan 25;250(2):661-7.
The interaction between the catalytic subunit (c3) and the regulatory subunit (r2) of aspartate transcarbamylase from Escherichia coli was studied by measuring the reversible formation of the c3r6 complex as a function of r2 concentration. Conversion to the native enzyme was prevented by using a very low concentration of c2 (40 ng per ml) in the presence of bovine serum albumin. A simple hyperbolic r2 saturation curve was obtained suggesting the presence of only one kind of c:r domain. From the association constant for the formation of c3r6, the free energy of c:r interaction can be estimated to be about -10 Cal per mole. Neither CTP nor ATP appears to affect the strength of c:r interaction in this complex. Succinate in the presence of carbamyl phosphate promotes tighter binding. At higher concentration of c3 and nonsaturating levels of r2, conversion to the native enzyme (c3r6) takes place. This renaturation process is second order with respect to the concentration of c3 and is virtually irreversible. Renaturation is inhibited by saturating levels of r2 and to some extent by both CTP and ATP. The effect of ligands on c:r interactions reported here may have significance in the allosteric mechanism of the native enzyme.
通过测量c3r6复合物的可逆形成作为r2浓度的函数，研究了来自大肠杆菌的天冬氨酸转氨甲酰酶的催化亚基（c3）和调节亚基（r2）之间的相互作用。在牛血清白蛋白存在下，通过使用非常低浓度的c2（每毫升40纳克）来防止转化为天然酶。获得了一条简单的双曲线r2饱和曲线，表明仅存在一种c:r结构域。根据c3r6形成的缔合常数，c:r相互作用的自由能估计约为每摩尔-10卡路里。在该复合物中，CTP和ATP似乎都不影响c:r相互作用的强度。在氨基甲酰磷酸存在下的琥珀酸盐促进更紧密的结合。在较高浓度的c3和不饱和水平的r2下，会发生向天然酶（c3r6）的转化。这种复性过程相对于c3的浓度是二级的，并且实际上是不可逆的。r2的饱和水平会抑制复性，CTP和ATP在一定程度上也会抑制复性。此处报道的配体对c:r相互作用的影响可能在天然酶的变构机制中具有重要意义。
