Kovina Marina V, De Kok Aart, Sevostyanova Irina A, Khailova Ludmila S, Belkina Natalya V, Kochetov German A
A.N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia.
Proteins. 2004 Aug 1;56(2):338-45. doi: 10.1002/prot.20115.
New and previously published data on a variety of ThDP-dependent enzymes such as baker's yeast transketolase, yeast pyruvate decarboxylase and pyruvate dehydrogenase from pigeon breast muscle, bovine heart, bovine kidney, Neisseria meningitidis and E. coli show their spectral sensitivity to ThDP binding. Although ThDP-induced spectral changes are different for different enzymes, their universal origin is suggested as being caused by the intrinsic absorption of the pyrimidine ring of ThDP, bound in different tautomeric forms with different enzymes. Non-enzymatic models with pyrimidine-like compounds indicate that the specific protein environment of the aminopyrimidine ring of ThDP determines its tautomeric form and therefore the changeable features of the inducible effect. A polar environment causes the prevalence of the aminopyrimidine tautomeric form (short wavelength region is affected). For stabilization of the iminopyrimidine tautomeric form (both short- and long-wavelength regions are affected) two factors appear essential: (i) a nonpolar environment and (ii) a conservative carboxyl group of a specific glutamate residue interacting with the N1' atom of the aminopyrimidine ring. The two types of optical effect depend in a different way upon the pH, in full accordance with the hypothesis tested. From these studies it is concluded that the inducible optical rotation results from interaction of the aminopyrimidine ring with its asymmetric environment and is defined by the protonation state of N1' and the 4'-nitrogen.
关于多种硫胺素二磷酸（ThDP）依赖性酶的新数据以及先前发表的数据，如面包酵母转酮醇酶、酵母丙酮酸脱羧酶和来自鸽胸肌、牛心脏、牛肾脏、脑膜炎奈瑟菌和大肠杆菌的丙酮酸脱氢酶，显示了它们对ThDP结合的光谱敏感性。尽管不同酶的ThDP诱导光谱变化不同，但它们的普遍来源被认为是由以不同互变异构形式与不同酶结合的ThDP嘧啶环的固有吸收引起的。含有嘧啶类化合物的非酶模型表明，ThDP氨基嘧啶环的特定蛋白质环境决定了其互变异构形式，从而决定了诱导效应的可变特征。极性环境导致氨基嘧啶互变异构形式占优势（短波长区域受到影响）。对于亚氨基嘧啶互变异构形式的稳定（短波长和长波长区域均受到影响），两个因素似乎至关重要：（i）非极性环境和（ii）与氨基嘧啶环的N1'原子相互作用的特定谷氨酸残基的保守羧基。这两种光学效应以不同方式依赖于pH，这与所测试的假设完全一致。从这些研究得出的结论是，诱导旋光性是由氨基嘧啶环与其不对称环境的相互作用产生的，并由N1'和4'-氮的质子化状态决定。
