Lai J C, Cooper A J
J Neurochem. 1986 Nov;47(5):1376-86. doi: 10.1111/j.1471-4159.1986.tb00768.x.
The substrate and cofactor requirements and some kinetic properties of the alpha-ketoglutarate dehydrogenase complex (KGDHC; EC 1.2.4.2, EC 2.3.1.61, and EC 1.6.4.3) in purified rat brain mitochondria were studied. Brain mitochondrial KGDHC showed absolute requirement for alpha-ketoglutarate, CoA and NAD, and only partial requirement for added thiamine pyrophosphate, but no requirement for Mg2+ under the assay conditions employed in this study. The pH optimum was between 7.2 and 7.4, but, at pH values below 7.0 or above 7.8, KGDHC activity decreased markedly. KGDHC activity in various brain regions followed the rank order: cerebral cortex greater than cerebellum greater than or equal to midbrain greater than striatum = hippocampus greater than hypothalamus greater than pons and medulla greater than olfactory bulb. Significant inhibition of brain mitochondrial KGDHC was noted at pathological concentrations of ammonia (0.2-2 mM). However, the purified bovine heart KGDHC and KGDHC activity in isolated rat heart mitochondria were much less sensitive to inhibition. At 5 mM both beta-methylene-D,L-aspartate and D,L-vinylglycine (inhibitors of cerebral glucose oxidation) inhibited the purified heart but not the brain mitochondrial enzyme complex. At approximately 10 microM, calcium slightly stimulated (by 10-15%) the brain mitochondrial KGDHC. At concentrations above 100 microM, calcium (IC50 = 1 mM) inhibited both brain mitochondrial and purified heart KGDHC. The present results suggest that some of the kinetic properties of the rat brain mitochondrial KGDHC differ from those of the purified bovine heart and rat heart mitochondrial enzyme complexes. They also suggest that the inhibition of KGDHC by ammonia and the consequent effect on the citric acid cycle fluxes may be of pathophysiological and/or pathogenetic importance in hyperammonemia and in diseases (e.g., hepatic encephalopathy, inborn errors of urea metabolism, Reye's syndrome) where hyperammonemia is a consistent feature. Brain accumulation of calcium occurs in a number of pathological conditions. Therefore, it is possible that such a calcium accumulation may have a deleterious effect on KGDHC activity.
对纯化的大鼠脑线粒体中α-酮戊二酸脱氢酶复合体（KGDHC；EC 1.2.4.2、EC 2.3.1.61和EC 1.6.4.3）的底物、辅因子需求及一些动力学特性进行了研究。脑线粒体KGDHC对α-酮戊二酸、辅酶A和NAD有绝对需求，对添加的硫胺焦磷酸只有部分需求，但在本研究采用的测定条件下对Mg2+无需求。最适pH在7.2至7.4之间，但在pH值低于7.0或高于7.8时，KGDHC活性显著下降。不同脑区的KGDHC活性顺序为：大脑皮层＞小脑≥中脑＞纹状体 = 海马＞下丘脑＞脑桥和延髓＞嗅球。在病理性氨浓度（0.2 - 2 mM）下，观察到脑线粒体KGDHC受到显著抑制。然而，纯化的牛心KGDHC和分离的大鼠心脏线粒体中的KGDHC活性对抑制的敏感性要低得多。在5 mM时，β-亚甲基-D,L-天冬氨酸和D,L-乙烯基甘氨酸（脑葡萄糖氧化抑制剂）均抑制纯化的心脏但不抑制脑线粒体酶复合体。在约10 μM时，钙轻微刺激（10 - 15%）脑线粒体KGDHC。在浓度高于100 μM时，钙（IC50 = 1 mM）抑制脑线粒体和纯化的心脏KGDHC。目前的结果表明，大鼠脑线粒体KGDHC的一些动力学特性与纯化的牛心和大鼠心脏线粒体酶复合体不同。它们还表明，氨对KGDHC的抑制以及随后对柠檬酸循环通量的影响在高氨血症以及高氨血症为一致特征的疾病（如肝性脑病、尿素代谢先天性缺陷、瑞氏综合征）中可能具有病理生理和/或致病重要性。在许多病理状况下会出现脑钙蓄积。因此，这种钙蓄积可能对KGDHC活性产生有害影响。
