BRAIN GLYCOGEN PHOSPHORYLASE IN HYPOXIC-ISCHEMIC EVENTS

缺氧缺血事件中的脑糖原磷酸化酶

• 批准号: 3407397
• 负责人: FREDRIC A GORIN
• 金额: $ 6.05万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-09-01 至 1987-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3407397
• 关键词: X ray crystallography; adenosine monophosphate; adenosine triphosphate; allosteric site; bioenergetics; brain cell; brain metabolism; brain regulatory center; central nervous system; cerebral cortex; cerebral ischemia /hypoxia; complementary DNA; convulsions; genetic manipulation; glia; glycogenolysis; hypoglycemia; isozymes; messenger RNA; molecular cloning; neurochemistry; nucleic acid sequence; phosphorylases; radioassay; striated muscles

Brain glycogenolysis plays a critical role in meeting the acute energy requirements of the central nervous system during hypoxic-ischemic events, hypoglycemia, and seizures. Recent evidence suggests that brain glycogen phosphorylase has important regulatory differences from the isoenzymes expressed by skeletal muscle and liver. These regulatory differences, in particular inhibition of brain phosphorylase by glucose, has important implications into understanding control of brain glycogenolysis during acute alterations in cellular metabolism of brain tissue. We propose to identify and determine the amino sequence of brain glycogen phosphorylase in rabbit using recombinant DNA techniques. Our laboratory has determined the x-ray crystallographic structure of rabbit muscle phosphorylase a to 2.1 Angstrom resolution. We have co-crystallized regulatory ligands with the enzyme, identified five regulatory sites for this enzyme, and have elucidated many of the molecular mechanisms governing their allosteric control. We will compare the amino acid sequence of rabbit brain phosphorylase with the known sequence and tertiary structure of the rabbit muscle isoenzyme. This should allow us by comparison to account for the maintenance or alteration of regulatory sites for the brain isoenzyme. We will partially purify brain phosphorylase in rabbit from the other phosphorylase isoenzymes and examine the regulatory properties of purine compounds, AMP, ATP, as well as selective gluconeogenic substrates. DNA probes encoding for fragments of brain and skeletal muscle glycogen phosphorylase will be used for in situ hybridization to regions of the neocortex to localize these isoenzymes to neuronal and glial elements within the central nervous system.

脑糖基溶解在满足急性能量方面起着关键作用 缺氧 - 缺血事件期间中枢神经系统的要求， 低血糖和癫痫发作。 最近的证据表明脑糖原 磷酸化酶具有与同工酶的重要调节差异 由骨骼肌和肝脏表达。 这些监管差异， 葡萄糖对脑磷酸化酶的特别抑制作用具有重要 在理解控制脑糖原分解的影响 脑组织细胞代谢的急性改变。 我们建议识别和确定脑糖原的氨基序列 使用重组DNA技术在兔中的磷酸化酶。 我们的实验室 已经确定了兔肌肉的X射线晶体结构 磷酸化酶A至2.1 Angstrom分辨率。 我们共结晶了 具有酶的调节配体，确定了五个调节位点 这种酶，并阐明了许多分子机制 他们的变构控制。 我们将比较 具有已知序列和第三纪结构的兔脑磷酸化酶 兔肌肉同工酶的。 相比之下，这应该使我们 说明大脑调节地点的维护或改变 同工酶。 我们将部分净化兔子中的脑磷酸化酶 其他磷酸化酶同工酶并检查 嘌呤化合物，AMP，ATP以及选择性糖原底物。 DNA探针编码大脑碎片和骨骼肌糖原的探针 磷酸化酶将用于原位杂交与 将这些同工酶定位于神经元和神经胶质元素的新皮层 在中枢神经系统中。