Ligand extraction properties of the GM2 activator protein and its interactions with lipid vesicles.
Ligand extraction properties of the GM2 activator protein and its interactions with lipid vesicles.
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GM2 激活蛋白的配体提取特性及其与脂质囊泡的相互作用。
DOI：
10.1016/j.bpj.2009.03.065
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发表时间：
2009
影响因子：
3.4
通讯作者：
Fanucci,GailE
中科院分区：
文献类型：
--
作者：
Ran,Yong;Fanucci,GailE
GM2激活蛋白（GM2AP）是细胞溶酶体区室中水解酶将GM2酶转化为GM3所需的辅助蛋白。本研究通过糖负载囊泡沉降和凝胶过滤实验研究了GM2AP与脂质囊泡的相互作用，并表征了pH和脂质组成对膜结合和脂质提取的影响。沉淀实验允许对溶液和双层表面的蛋白质百分比进行简单的定量分析，并对溶液中保留的蛋白质：脂质复合物进行详细分析。最佳的结合和配体提取被发现在pH为4.8时，无论脂质组成如何，<15%的蛋白质保持表面相关。除了提取GM2外，我们发现GM2AP很容易从囊泡中提取丹酰头基团标记的脂质以及其他磷脂。GM2AP从囊泡中提取丹酰二氢聚乙烯的能力受pH和特定配体GM2的影响。虽然独特的内体脂质，二磷酸单酰基甘油，不需要提取配体，但当胆固醇存在于囊泡中时，它确实提高了GM2的提取效率。
The GM2 activator protein (GM2AP) is an accessory protein required for the enzymatic conversion of GM2 to GM3 by hydrolases in the lysosomal compartments of cells. Here, GM2AP interactions with lipid vesicles are investigated by sucrose-loaded vesicle sedimentation and gel filtration assays, and the effects of pH and lipid composition on membrane binding and lipid extraction are characterized. The sedimentation experiments allow for facile quantification of the percentage of protein in solution and on the bilayer surface, with detailed analysis of the protein:lipid complex that remains in solution. Optimum binding and ligand extraction is found for pH 4.8 where <15% of the protein remains surface associated regardless of the lipid composition. In addition to extracting GM2, we find that GM2AP readily extracts dansyl-headgroup-labeled lipids as well as other phospholipids from vesicles. The ability of GM2AP to extract dansyl-DHPE from vesicles is altered by pH and the specific ligand GM2. Although the unique endosomal lipid, bis(monoacylglycero)phosphate, is not required for ligand extraction, it does enhance the extraction efficiency of GM2 when cholesterol is present in the vesicles.