BRAIN PYRUVATE DEHYDROGENASE: NEUROBIOLOGY AND REGULATI

脑丙酮酸脱氢酶：神经生物学和调节

• 批准号: 3405806
• 负责人: KWAN-FU REX SHEU
• 金额: $ 6.2万
• 依托单位: BURKE REHABILITATION CTR (WHITE PLNS,NY)
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-02-01 至 1989-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3405806
• 关键词: aging; brain metabolism; calcium metabolism; gel electrophoresis; genetic translation; immunochemistry; ionophores; mitochondria; neurochemistry; oxidation; phosphorylation; pyruvate dehydrogenase; radioimmunoassay

The brain energy metabolism is met primarily in mitochondria by oxidative metabolism of pyruvate, which is regulated by pyruvate dehydrogenase complex (PDHC). The PDHC is controlled by a kinase-phosphatase mediated phosphorylation (inactivation) and dephosphorylation (activation) process. Deficits of PDHC have been found in an increasing number of neurological disorders, including Alzheimer's disease. Changes of phosphorylation or activation state of PDHC have also been found to accompany many physiological, pharmacological and behavioral manipulations that give rise to altered brain functions. These changes of PDHC may reflect changes of pyruvate oxidation or Ca sequestration by mitochondria. The molecular events underlying these changes are unclear, however. To this end, the relationship among PDHC regulatory behavior, pyruvate ovidation and mitochondrial Ca sequestration will be delineated in this proposition. The mitochondrial Ca-uptake process is distinct from the effect of intramitochondrially accumulated Ca. The effects of these two processes on pyruvate oxidation and PDHC phosphorylation will be evaluated, using mitochondria treated with ionophore A23187 in Ca-EGTA buffer and mitochonria pre-loaded or depleted with Ca, respectively. With respect to enzyme protein, PDHC may be regulated by phosphorylation at multiple sites and by protein translation during postnatal development. The significance of multiple phosphorylation will be evaluated by comparing the phosphate profile of the phosphopeptide generated by immunoprecipitation and 2-dimensional gel electrophoresis. The enzyme induction mechanism will be tested by radioimmunoassays and in vivo translation assays of the PDHC-peptides. Aging decreases pyruvate oxidation and Ca metabolism. To further test the hypothesis that mitochondrial Ca sequestration is also imparied in aging, the Ca buffering capacity, and Ca uptake and its coupling to pyruvate oxidation and possible link to PDHC phosphorylation will be comared in mitochondria from different age groups.

大脑的能量代谢主要在线粒体中通过氧化来满足 丙酮酸代谢，受丙酮酸脱氢酶调节 复合物（PDHC）。 PDHC 受激酶磷酸酶介导的控制 磷酸化（失活）和去磷酸化（激活）过程。 越来越多的神经系统疾病患者发现 PDHC 缺陷 疾病，包括阿尔茨海默病。 磷酸化的变化或 PDHC 的激活状态也被发现伴随着许多 生理、药理学和行为操纵，从而产生 改变大脑功能。 PDHC 的这些变化可能反映了 线粒体的丙酮酸氧化或钙隔离。 分子 然而，这些变化背后的事件尚不清楚。 为此， PDHC 调节行为、丙酮酸产卵和 本命题将描述线粒体钙隔离。 线粒体钙吸收过程不同于 线粒体内积累的 Ca。 这两个过程的影响 将使用以下方法评估丙酮酸氧化和 PDHC 磷酸化 用 Ca-EGTA 缓冲液中的离子载体 A23187 处理线粒体， 线粒体分别预载或耗尽 Ca。 就酶蛋白而言，PDHC可能通过磷酸化来调节 产后发育过程中的多个位点和蛋白质翻译。 通过比较来评估多重磷酸化的重要性 产生的磷酸肽的磷酸谱 免疫沉淀和二维凝胶电泳。 酶 诱导机制将通过放射免疫分析和体内测试进行测试 PDHC 肽的翻译测定。 衰老会降低丙酮酸氧化和钙代谢。 为了进一步测试 假设线粒体钙隔离也会因衰老而受到损害， Ca 缓冲能力、Ca 吸收及其与丙酮酸的偶联 氧化和可能与 PDHC 磷酸化的联系将在 来自不同年龄组的线粒体。