FATTY ACID ACYLATION OF MYELIN GLYCOPROTEIN

髓磷脂糖蛋白的脂肪酸酰化

• 批准号: 2266781
• 负责人: OSCAR A BIZZOZERO
• 金额: $ 9.49万
• 依托单位: UNIVERSITY OF NEW MEXICO
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-07-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2266781
• 关键词: acyl coA; acyl group; acylation; acyltransferase; cell free system; covalent bond; enzyme substrate; fatty acid metabolism; glycosylation; high performance liquid chromatography; laboratory mouse; laboratory rat; mass spectrometry; myelin; myelin glycoprotein; myelination; neurochemistry; neurogenesis; nucleic acid sequence; palmitates; peptidases; peptides; peripheral nervous system; phosphorylation; physical separation; protein biosynthesis; tritium

Our long term goal is to understand the mechanisms that control the formation and maintenance of the CNS and PNS myelin, PO, the major protein of PNS myelin, participates in the formation and compaction of the myelin lamellae. PO is esterified with long chain fatty acids, as is PLP, the major protein of CNS myelin. We hypothesize that acylation plays an important role in the process of myelin synthesis and maintenance. The present studies are aimed at determining fundamental aspects of the biology of PO acylation. Our specific aims are: 1. To study the metabolism of the fatty acid bound to PO using a nerve slice system. Double-label pulse and pulse-chase experiments, combined with subcellular fractionation, will be used to identify the membranes in which acylation of PO takes place. We will establish the relationship between acylation and the other PO posttranslational modifications by using inhibitors of protein N-glycosylation and phosphorylation. We will also correlate the extent of PO synthesis and acylation during nerve development to gain insights on the role of this modification. Our preliminary results indicate that acylation occurs only on newly-synthesized but also on a pre- existing PO, suggesting that acylation could also relate to myelin maintenance. 2. To determine the acylation site on the PO molecule. PO will be labeled with [3H] palmitic acid, digested with several proteases and the acyl- peptides isolated and sequenced. 3. To develop a cell-free system using deacylated PO and labeled acyl-CoA as substrates. We will carry out a detailed characterization of the acylating enzyme, localize the subcellular site of cylation unequivocally, isolate the acyltransferase and determine its protein substrate specificity, and determine the developmental and evolutionary changes of this enzyme. The studies proposed in this application will provide basic knowledge on the biology of PO acylation. Moreover, they will contribute to gain insights into normal myelination processes and ultimately to understand the pathophysiology of human peripheral nerve disorders.

我们的长期目标是了解控制 CNS和PNS髓鞘的形成和维持，PO是主要蛋白质 PNS髓鞘的形成，参与髓鞘的形成和致密化 悲伤。 PO被长链脂肪酸酯化，PLP也是如此， CNS髓鞘的主要蛋白质。 我们假设酰化作用是 在髓鞘合成和维持过程中起重要作用。 的 目前的研究旨在确定生物学的基本方面， PO酰化。 我们的具体目标是： 1. 使用神经研究与PO结合的脂肪酸的代谢 切片系统 双标记脉冲和脉冲追踪实验，结合 与亚细胞分级分离，将被用来确定膜， 其中PO发生酰化。 我们将建立关系 酰化和其他PO翻译后修饰之间的关系， 蛋白质N-糖基化和磷酸化的抑制剂。 我们还将 神经发育过程中PO合成和酰化程度的相关性 来了解这种改变的作用。 我们的初步结果 表明酰化作用不仅发生在新合成的上，而且也发生在预合成的上， 存在PO，表明酰化也可能与髓鞘有关 上维护 2. 确定PO分子上的酰化位点。 将标记PO 与[3 H]棕榈酸，用几种蛋白酶消化和酰基- 肽分离和测序。 3. 使用脱酰PO和标记的酰基-CoA开发无细胞系统 作为底物。 我们将进行详细的表征， 酰化酶，明确地定位亚细胞的酰化位点， 分离酰基转移酶并确定其蛋白底物 特异性，并确定发展和进化的变化， 这种酶。 本申请中提出的研究将提供以下方面的基本知识 PO酰化的生物学。 此外，他们将有助于获得 了解正常的髓鞘形成过程，并最终了解 人类周围神经疾病的病理生理学。