Heink Anna, Davidson W Sean, Swertfeger Debi K, Lu L Jason, Shah Amy S
§Center for Lipid and Arteriosclerosis Science, Department of Pathology and Laboratory Medicine, University of Cincinnati, 2120 East Galbraith Road, Cincinnati, Ohio 45237-0507, United States.
J Proteome Res. 2015 Jul 2;14(7):2943-50. doi: 10.1021/acs.jproteome.5b00270. Epub 2015 Jun 12.
We sought to develop a new method to more efficiently analyze lipid-bound proteins by mass spectrometry using a combination of a lipid removal agent (LRA) that selectively targets lipid-bound proteins and a mass spectrometry compatible detergent, anionic acid labile surfactant (AALS), that is capable of eluting proteins off the LRA. This method was compared to established methods that use the lipid removal agent alone and straight proteomic analysis of human plasma after organic solvent delipidation (OSD). Plasma from healthy individuals was separated by gel filtration chromatography and prepared for mass spectrometry analysis by each of the described methods. The addition of AALS to LRA increased the overall number of proteins detected in both the high and low density lipoprotein size range, the number of peptide counts for each protein, and the overall sequence coverage. Organic solvent delipidation detected the most proteins, though with some decrease in overall protein detection and sequence coverage due to the presence of nonlipid-bound proteins. The use of LRA allows for selection and analysis of lipid-bound proteins. The addition of a mass spectrometry compatible detergent improved detection of lipid-bound proteins from human plasma using LRA.
我们试图开发一种新方法，通过质谱法更有效地分析脂质结合蛋白，该方法结合了一种选择性靶向脂质结合蛋白的脂质去除剂（LRA）和一种与质谱兼容的去污剂——阴离子酸不稳定表面活性剂（AALS），后者能够将蛋白从LRA上洗脱下来。将该方法与单独使用脂质去除剂的既定方法以及有机溶剂脱脂（OSD）后人血浆的直接蛋白质组分析方法进行了比较。通过凝胶过滤色谱法分离健康个体的血浆，并采用上述每种方法进行质谱分析准备。在LRA中添加AALS增加了在高密度和低密度脂蛋白大小范围内检测到的蛋白质总数、每种蛋白质的肽计数以及总体序列覆盖率。有机溶剂脱脂检测到的蛋白质最多，不过由于存在非脂质结合蛋白，总体蛋白质检测和序列覆盖率有所下降。使用LRA可以选择和分析脂质结合蛋白。添加与质谱兼容的去污剂可改善使用LRA从人血浆中检测脂质结合蛋白的效果。
