Pyridoxal 5′-phoshate Schiff base in Citrobacter freundii tyrosinephenol-lyase -: Ionic and tautomeric equilibria
Pyridoxal 5′-phoshate Schiff base in Citrobacter freundii tyrosinephenol-lyase -: Ionic and tautomeric equilibria
复制标题
DOI：
10.1046/j.1432-1327.2000.01185.x
复制
发表时间：
2000-03-01
期刊：
影响因子：
--
通讯作者：
Demidkina, TV
中科院分区：
文献类型：
--
作者：
Bazhulina, NP;Morozov, YV;Demidkina, TV
在激活阳离子K+和阳离子抑制剂Na+存在的情况下，研究了酪氨酸苯酚裂合酶内部希夫碱的光谱性质。在pH 6.5-8.7范围内，在K+存在下，记录了全酶的吸收光谱。在该pH范围内没有发现辅酶发色团的明显pK（a）值，表明内部席夫碱在从pH 6.5到8.7时不改变其离子形式。为了确定酪氨酸酚裂解酶中席夫碱的离子状态和互变异构体组成，利用对数正态分布曲线分析了吸收光谱和圆二色性光谱。内部席夫碱的主要形式是具有质子化的吡啶鎓和醛亚胺氮原子和去质子化的3 '-羟基的席夫碱，即酮烯胺。该形式与其烯醇亚胺互变异构体处于质子转移平衡。内部醛亚胺离子形式在用Na+置换K+时改变。这种置换导致吡哆醛-P的吡啶鎓氮的pK（a）值显著降低。
Spectral properties of the internal Schiff base in tyrosine phenol-lyase have been investigated in the presence of an activating cation K+ and a cation-inhibitor Na+. The holoenzyme absorption spectra in the pH range 6.5-8.7 were recorded in the presence of K+. No apparent pK(a) value of the coenzyme chromophore was found in this pH range, indicating that the internal Schiff base does not change its ionic form on going from pH 6.5 to 8.7. To determine the ionic state and tautomeric composition of the Schiff base in tyrosine phenol-lyase, the absorption and circular dichroism spectra were analyzed using lognormal distribution curves. The predominant form of the internal Schiff base is that with protonated pyridinium and aldimine nitrogen atoms and deprotonated 3'-hydroxy group, i.e. the ketoenamine. This form is in prototropic equilibrium with its enolimine tautomer. The internal aldimine ionic form is changed upon replacement of K+ with Na+. This replacement leads to a significant decrease in the pK(a) value of pyridinium nitrogen of the pyridoxal-P.