Cho S W, Lee J E
Department of Biochemistry, College of Medicine, University of Ulsan, Seoul, Korea.
Biochimie. 1996;78(10):817-21. doi: 10.1016/s0300-9084(97)84333-7.
Two soluble forms of brain glutamate dehydrogenase isoproteins were inactivated by pyridoxal 5'-phosphate. Restoration of catalytic activity can be accomplished by dialysis and addition of an excess of cysteine or lysine. Spectral evidence is presented to indicate that the inactivation proceeds through Schiff base formation with amino groups of the enzyme. Inactivation became irreversible after reduction with NaBH4 and the NaBH4-reduced enzyme showed a characteristic absorption peak at 325 nm. Using spectral titration at 325 nm, the stoichiometry was 2 mol/mol of GDH subunit without protection and 1 mol/mol with protection, indicating the complete masking of one mol of lysine. The results with analogs of pyridoxal 5'-phosphate show that the aldehyde group, but not the phosphate group, is required for efficient inactivation.
两种可溶性形式的脑谷氨酸脱氢酶同工蛋白被磷酸吡哆醛失活。通过透析并添加过量的半胱氨酸或赖氨酸可以恢复催化活性。光谱证据表明，失活是通过与酶的氨基形成席夫碱进行的。用硼氢化钠还原后，失活变得不可逆，并且硼氢化钠还原的酶在325nm处显示出特征吸收峰。使用325nm处的光谱滴定法，化学计量比在无保护时为每摩尔GDH亚基2摩尔，有保护时为每摩尔1摩尔，表明一摩尔赖氨酸被完全掩盖。磷酸吡哆醛类似物的结果表明，有效失活需要醛基而非磷酸基团。
