FATTY ACID ACYLATION OF MYELIN GLYCOPROTEIN

我们的长期目标是了解控制 中枢神经系统和三叉神经节髓鞘主要蛋白PO的形成和维持 PNS髓磷脂，参与髓鞘的形成和紧实 薄片。PO与长链脂肪酸一起酯化，PLP也是如此， 中枢神经系统髓鞘的主要蛋白质。我们假设酰化作用是一种 在髓鞘合成和维持过程中起重要作用。这个 目前的研究旨在确定生物学的基本方面。 PO酰化反应的影响。我们的具体目标是： 1.用神经研究PO结合的脂肪酸代谢 切片系统。双标记脉冲和脉冲追逐相结合的实验 通过亚细胞分离，将被用来鉴定 PO发生了哪种酰化反应。我们将建立这种关系 在酰化和其他PO翻译后修饰之间使用 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 以深入了解这一修改的作用。我们的初步结果 表明酰化反应只发生在新合成的化合物上，也发生在预合成的 存在PO，表明酰化也可能与髓鞘有关 维修。 2.确定PO分子上的酰化位点。PO将被贴上标签 用[~3H]棕榈酸，用几种酶消化，酰基- 多肽的分离和测序。 3.利用脱酰化PO和标记的酰基辅酶A建立无细胞体系 作为底物。我们将进行详细的特性描述 酰化酶，明确定位细胞周期的亚细胞位置， 酰基转移酶的分离及其蛋白质底物的测定 特异性，并决定了其发育和进化的变化 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，他们还将为收益做出贡献。 深入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。

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.