Proteomic Identification of PKC-Mediated Expression of 20E-Induced Protein in Drosophila melanogaster

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• 作者：Yaning Sun ; Shiheng An ; Vincent C. Henrich ; Xiaoping Sun ; Qisheng Song
• 刊名：Journal of Proteome Research
• 出版年：2008
• 出版时间：April 2008
• 年：2008
• 卷：7
• 期：4
• 页码：1784 - 1784
• 全文大小：122K
• 年卷期：v.7,no.4(April 2008)
• ISSN：1535-3907

文摘

Analysis of membrane proteins, particularly integral membrane proteins, still presents a great challenge due to their poor water solubility and low abundance though much effort has been devoted to the solubilization and enrichment of the protein class. In this paper, a two-phase, on-membrane digestion method was developed and applied in the analysis of rat liver membrane proteome. The two-phase system was constituted by mixing n-butanol and 25 mM NH<SUB>4HCO₃. Comparative experiments indicated that the proteins on membranes could be digested in the two-phase system more efficiently than in both 60% methanol and 25 mM NH₄HCO₃ solutions under the same conditions, thereby improving the identification of the membrane proteins. When the established two-phase system and CapLC-MS/MS was used to analyze rat liver membrane proteome, a total of 411 membrane proteins were identified, more than 80% of which were transmembrane proteins with 1–12 mapped transmembrane domains (TMDs). Because of its extraction and dissolution actions, the two-phase on-membrane digestion system we developed could efficiently improve the digestion and removal of adsorbed nonmembrane proteins, and remarkably increase the number and coverage of identified membrane proteins, particularly the transmembrane proteins. Using our procedure to identify a complementary protein set from all fractions of the two-phase system could achieve a higher coverage of the membrane proteome.