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Issue 5, 2010
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Improving the depth of coverage in membrane proteomic studies through the use of lipid-based protein immobilization technology in parallel with methanol-facilitated solubilisation

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Abstract

Lipid-based protein immobilization (LPI) technology is a platform recently developed to facilitate shotgun membrane proteomic studies based on a nanotechnology framework. Proteoliposomes are generated from a membraneprotein preparation. These proteoliposomes are immobilized onto an LPI chip and then subjected to proteolysis. The proteolytic peptides are then subjected to LC/MS analysis after fractionation by SCX chromatography. The focus of this study was to evaluate how the depth of coverage of the membrane proteome of a particular cell type varied as a function of the sample preparation method used. Human dermal fibroblasts (hDFs) and human bone marrow mesenchymal stem cells (BM-hMSCs) were subjected to membrane proteomic studies using two different sample preparation methods: LPI technology and methanol-facilitated solubilisation. The number of membraneproteins that could be identified from hDFs and BM-hMSCs using LC/MS was greater using LPI technology than it was using methanol-facilitated solubilisation. However, the number of membraneprotein identifications that could be made for both hDFs and BM-hMSCs increased by ∼50% when both sample preparation methods were used in parallel and the MS/MS data was convolved together. Therefore, LPI technology is a very useful technology for high-throughput shotgun membrane proteomic studies. However, in order to maximize the depth of membrane proteome coverage that can be attained for a given cell type, it is necessary to use multiple sample preparation methods in concert.

Graphical abstract: Improving the depth of coverage in membrane proteomic studies through the use of lipid-based protein immobilization technology in parallel with methanol-facilitated solubilisation

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Supplementary files

Article information


Submitted
20 Nov 2009
Accepted
09 Feb 2010
First published
23 Feb 2010

Anal. Methods, 2010,2, 539-545
Article type
Paper

Improving the depth of coverage in membrane proteomic studies through the use of lipid-based protein immobilization technology in parallel with methanol-facilitated solubilisation

N. D. Padliya, M. B. Bhatia, W. T. Hofgärtner and R. J. Hariri, Anal. Methods, 2010, 2, 539
DOI: 10.1039/B9AY00267G

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