Osborne H H, Hollaway M R
Biochem J. 1975 Oct;151(1):37-45. doi: 10.1042/bj1510037.
An investigation was made of changes in subunit interactions in glyceraldehyde 3-phosphate dehydrogenase on binding NAD+, NADH and other substrates by using the previously developed method of measurement of rates and extent of subunit exchange between the rabbit enzyme (R4), yeast enzyme (Y4) and rabbit-yeast hybrid (R2Y2) [Osborne & Hollaway (1974) Biochem. J. 143, 651-662]. The free energy of activation for the conversion of tetramer into dimer for the rabbit enzyme (R4 leads to 2R2) is increased by at least 12kJ/mol in the presence of NAD+. This increase is interpreted in terms of an NAD+-induced 'tightening' of the tetrameric structure probably involving increased interaction at the subunit interfaces across the QR plane of the molecule [see Buehner et al. (1974) J. Mol. Biol. 82, 563-585]. This tightening of the structure only occurs on binding the third NAD+ molecule to a given enzyme molecule. Conversely, binding of NADH causes a decrease in the free energy of activation for the R4 leads to 2R2 and Y4 leads to 2Y2 conversions by at least 10kJ/mol. This is interpreted as a NADH-induced 'loosening' of the structures arising from decreased interactions across the subunit interfaces involving the QR dissociation plane. In the presence of NADH the increase in the rate of subunit exchange is such that it is not possible to separate the hybrid from the other species if electrophoresis is carried out with NADH in the separation media. In the presence of a mixture of NADH and NAD+ the effect of NAD+ on subunit exchange is dominant. The results are discussed in terms of the known co-operativty between binding sites in glyceraldehyde 3-phosphate dehydrogenases.
利用先前开发的测量兔酶（R4）、酵母酶（Y4）和兔 - 酵母杂交酶（R2Y2）之间亚基交换速率和程度的方法，研究了甘油醛 - 3 - 磷酸脱氢酶在结合NAD⁺、NADH和其他底物时亚基相互作用的变化[奥斯本和霍拉韦（1974年），《生物化学杂志》143卷，651 - 662页]。在存在NAD⁺的情况下，兔酶（R4转变为2R2）四聚体转化为二聚体的活化自由能至少增加12kJ/mol。这种增加被解释为NAD⁺诱导的四聚体结构“收紧”，可能涉及分子QR平面上亚基界面处相互作用的增加[见比纳等人（1974年），《分子生物学杂志》82卷，563 - 585页]。这种结构的收紧仅在将第三个NAD⁺分子结合到给定酶分子上时发生。相反，NADH的结合导致R4转变为2R2和Y4转变为2Y2的活化自由能至少降低10kJ/mol。这被解释为NADH诱导的结构“松弛”，这是由于涉及QR解离平面的亚基界面间相互作用减少所致。在存在NADH的情况下，亚基交换速率的增加使得如果在分离介质中使用NADH进行电泳，就无法将杂交酶与其他物种分开。在存在NADH和NAD⁺混合物的情况下，NAD⁺对亚基交换的影响占主导地位。根据甘油醛 - 3 - 磷酸脱氢酶中结合位点之间已知的协同作用对结果进行了讨论。
