Venard R, Jallon J M, Fourcade A, Iwatsubo M
Eur J Biochem. 1975 Sep 15;57(2):371-8. doi: 10.1111/j.1432-1033.1975.tb02310.x.
Optical characteristics of enzyme-reduced coenzyme complexes of yeast NADP-specific glutamate dehydrogenase have been investigated in the presence and absence of product (L-glutamate) and in the presence or absence of phosphate. The phosphate effect, pointed out in a previous work, is found again: inorganic phosphate (Pi) destabilizes the binary complex (E - NADPH), the dissociation constant of which is equal to 14 muM, a value much higher than that determined in Tris-HCl buffer: Kd = 0.9 muM. Concerning the role of phosphate some assumptions are drawn up with respect to a similar behaviour of Pi toward yeast glutamate dehydrogenase and ADP toward the beef liver enzyme. In the same way, L-glutamate induces a stabilization of the binary complex; this latter effect is unchanged in the presence of phosphate, yet it is less marked than in the case of beef liver glutamate dehydrogenase. Protein fluorescence, nucleotide fluorescence and circular dichroism measurements allowed the determination of three identical and independent NADPH binding sites per hexameric active unit. In analogy with beef liver enzyme, it seems that yeast glutamate dehydrogenase is a good model to study anticooperativity in ligand binding.
在有或没有产物（L-谷氨酸）以及有或没有磷酸盐存在的情况下，对酵母NADP特异性谷氨酸脱氢酶的酶还原辅酶复合物的光学特性进行了研究。在之前的一项工作中指出的磷酸盐效应再次被发现：无机磷酸盐（Pi）会使二元复合物（E - NADPH）不稳定，其解离常数等于14 μM，该值远高于在Tris-HCl缓冲液中测定的值：Kd = 0.9 μM。关于磷酸盐的作用，针对Pi对酵母谷氨酸脱氢酶的类似行为以及ADP对牛肝酶的类似行为提出了一些假设。同样，L-谷氨酸会诱导二元复合物的稳定；在有磷酸盐存在的情况下，后一种效应没有变化，但比牛肝谷氨酸脱氢酶的情况不那么明显。蛋白质荧光、核苷酸荧光和圆二色性测量允许确定每个六聚体活性单元有三个相同且独立的NADPH结合位点。与牛肝酶类似，酵母谷氨酸脱氢酶似乎是研究配体结合中反协同作用的一个很好的模型。
