Evaluation of six sample preparation procedures for qualitative and quantitative proteomics analysis of milk fat globule membrane

文献类型： 外文期刊

第一作者： Yang, Yongxin

作者： Yang, Yongxin;Anderson, Elizabeth;Zhang, Sheng;Yang, Yongxin

作者机构：

关键词： Filter-aided sample preparation; In-solution digestion; Milk fat globule membrane; Suspension trapping

期刊名称：ELECTROPHORESIS （ 影响因子：3.535； 五年影响因子：2.731 ）

ISSN： 0173-0835

年卷期： 2018 年 39 卷 18 期

页码：

收录情况： SCI

摘要： Proteomic analysis of membrane proteins is challenged by the proteins solubility and detergent incompatibility with MS analysis. No single perfect protocol can be used to comprehensively characterize the proteome of membrane fraction. Here, we used cow milk fat globule membrane (MFGM) proteome analysis to assess six sample preparation procedures including one in-gel and five in-solution digestion approaches prior to LC-MS/MS analysis. The largest number of MFGM proteins were identified by suspension trapping (S-Trap) and filter-aided sample preparation (FASP) methods, followed by acetone precipitation without clean-up of tryptic peptides method. Protein identifications with highest average coverage was achieved by Chloroform/MeOH, in-gel and S-Trap methods. Most distinct proteins were identified by FASP method, followed by S-Trap. Analyses by Venn diagram, principal-component analysis, hierarchical clustering and the abundance ranking of quantitative proteins highlight differences in the MFGM fraction by the all sample preparation procedures. These results reveal the biased proteins/peptides loss occurred in each protocol. In this study, we found several novel proteins that were not observed previously by in-depth proteomics characterization of MFGM fraction in milk. Thus, a combination of multiple procedures with orthologous properties of sample preparation was demonstrated to improve the protein sequence coverage and expression level accuracy of membrane samples.

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