Membrane Binding Properties of the GM2 Activator Protein

GM2 激活蛋白的膜结合特性

• 批准号: 7075522
• 负责人: GAIL E FANUCCI
• 金额: $ 27.23万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-05-05 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7075522
• 关键词: binding proteins; conformation; electron spin resonance spectroscopy; gangliosides; glycosphingolipids; lipid bilayer membrane; lipid metabolism; lysosomes; membrane structure; protein binding; protein structure function

DESCRIPTION (provided by applicant): The GM2 Activator Protein (GM2AP) plays an essential role in the catabolism of GM2 in lysosomes. GM2 is a complex glycosphingolipid (GSL) that is a degradation intermediate in the breakdown of GM1 which is found in relatively high concentrations in the outer surface of plasma membranes in neuronal cells. GM2AP acts as a substrate specific co-factor by solubilizing GM2 from the intralysozomal vesicles and presenting the oligosaccharide moiety to beta-hexosaminidase A (HexA) for hydrolytic cleavage. Mutations in either HexA or GM2AP lead to the storage of GM2 within lysozomes and cell death. A well known example of this type of lysosomal storage disease is Tay-Sachs syndrome. The mechanism for how the GM2AP accessory protein regulates enzymatic degradation of GM2 as the first step in the GM2 lipid metabolism remains unclear. The proposed work will focus on elucidating the molecular details of this mechanism through site-directed spin labeling anomposition of late endosomes and intralysosomal vesicles are believed to alter the bilayer physical properties allowing GM2AP to extract GM2 from membranes. The current hypothesis is that in membranes of high curvature of neuronal lysozome composition, the binding is a transient event. Fluorescence based binding measurements will be used to test this hypothesis. Recently, crystal structures of the GM2AP bound to GM2, POPG and PAF were reported. The model of conformational changes predicted by X-ray crystallography data for specific and non-specific substrates will be tested by SDSL EPR experiments of GM2AP in solution where GM2 is extracted from micelles, as well as from lipid bilayers. Currently, the GM2AP has been expressed in E.coli, insect cells and in yeast. The effects of glycosylation on the membrane binding properties will also be examined. The general relevance of this research is that congenital mutations in GM2AP or HexA result in lysozomal storage diseases. The GM2-gangliosidoses (including Tay-Sachs disease) are a group of inherited disorders that result from defects in the catabolism of GM2. In fact, a variety of lysosomal storage diseases (Pompe, Fabry and the gangliosidoses) result from aberrations in lipid metabolism. A more detailed understanding of this critical first step in GM2 catabolism may lead to developments in drug or gene therapies for these fatal diseases.

描述（由申请人提供）：GM2激活蛋白（GM2AP）在GM2的分解代谢中起着至关重要的作用。 GM2是一种复杂的糖磷脂（GSL），是GM1分解中的降解中间体，在神经元细胞中质膜外表面相对较高的浓度中发现。 GM2AP通过将GM2溶解在侧面囊泡囊泡中，并将寡糖部分向β-己糖胺酶A（HEXA）呈现用于水解裂解的寡糖部分，从而充当底物特异性co因子。 HEXA或GM2AP中的突变导致GM2在溶菌酶和细胞死亡中的储存。 Tay-Sachs综合征是这种类型的溶酶体储存疾病的众所周知的例子。 GM2AP辅助蛋白如何调节GM2的酶促降解作为GM2脂质代谢的第一步的机制尚不清楚。拟议的工作将着重于阐明该机制的分子细节，该机制通过定位的旋转标记后期内体和肌体内囊泡的分子构成可以改变双层物理特性，从而允许GM2AP从膜上提取GM2。当前的假设是，在神经元溶菌酶组成的高曲率的膜中，结合是瞬态事件。基于荧光的结合测量将用于检验该假设。最近，报道了与GM2，POPG和PAF结合的GM2AP的晶体结构。 X射线晶体学数据对特定和非特异性底物预测的构象变化模型将通过GM2AP的SDSL EPR实验在溶液中测试，其中从胶束中提取GM2，以及从脂质双层中提取GM2。目前，GM2AP在大肠杆菌，昆虫细胞和酵母中都表达。还将检查糖基化对膜结合特性的影响。这项研究的一般意义是GM2AP或HEXA中的先天性突变会导致溶菌酶储存疾病。 GM2 - 长蛋白酶（包括Tay-Sachs疾病）是一组遗传性疾病，这些疾病是由于GM2分解代谢缺陷而导致的。实际上，脂质代谢的畸变导致了多种溶酶体储存疾病（蓬松，法布里和神经节蛋白）。对GM2分解代谢的这一关键第一步的更详细的了解可能会导致这些致命疾病的药物或基因疗法的发展。