Graham L D, Griffin T O, Beatty R E, McCarthy A D, Tipton K F
Biochim Biophys Acta. 1985 Apr 29;828(3):266-9. doi: 10.1016/0167-4838(85)90307-3.
In the presence of glutaric acid, N2,N2'-adipodihydrazido-bis(N6-carbonylmethyl-NAD+)(bis-NAD+ ) forms cross-links between molecules of glutamate dehydrogenase, resulting in precipitation. The dependence of this process on bis-NAD+ and enzyme concentration has been investigated. This procedure has been shown to be effective in the purification of glutamate dehydrogenase from rat and ox liver, and a procedure is presented in which this affinity precipitation procedure is used instead of the affinity chromatography used in an earlier method (McCarthy, A.D., Walker, J.M. and Tipton, K.F. (1980) Biochem. J. 191, 605-611). The ox liver enzyme prepared in this way had not suffered the limited proteolysis that occurs during the preparation of the enzyme by other commonly used procedures. After the purified enzyme had been denatured by treatment with urea, guanidine hydrochloride, or low pH, no recovery of activity could be demonstrated following dilution or, in the last case, dialysis.
在戊二酸存在的情况下，N2,N2'-己二酰二肼基双（N6-羰基甲基-NAD+）（双-NAD+）在谷氨酸脱氢酶分子之间形成交联，导致沉淀。研究了该过程对双-NAD+和酶浓度的依赖性。已证明该方法在从大鼠和牛肝中纯化谷氨酸脱氢酶方面是有效的，并且给出了一种方法，其中使用这种亲和沉淀法代替早期方法中使用的亲和色谱法（麦卡锡，A.D.，沃克，J.M.和蒂普顿，K.F.（1980年）《生物化学杂志》191，605 - 611）。以这种方式制备的牛肝酶没有受到其他常用方法制备酶过程中发生的有限蛋白水解作用。在用尿素、盐酸胍或低pH处理使纯化的酶变性后，稀释或（在最后一种情况下）透析后均未显示出活性恢复。
