Journal article
Triton X-114 phase separation in the isolation and purification of mouse liver microsomal membrane proteins
RA Mathias, YS Chen, EA Kapp, DW Greening, S Mathivanan, RJ Simpson
Methods | ACADEMIC PRESS INC ELSEVIER SCIENCE | Published : 2011
Abstract
Integral membrane proteins (IMPs) mediate several cellular functions including cell adhesion, ion and nutrient transport, and cell signalling. IMPs are typically hard to isolate and purify due to their hydrophobic nature and low cellular abundance, however, microsomes are small lipid vesicles rich in IMPs, which form spontaneously when cells are mechanically disrupted. In this study, we have employed mouse liver microsomes as a model for optimising a method for IMP isolation and characterisation. Microsomes were collected by differential centrifugation, purified with sodium carbonate, and subjected to GeLC-MS/MS analysis. A total of 1124 proteins were identified in the microsome fraction, wi..
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Awarded by Australian Cancer Research Foundation
Funding Acknowledgements
This work was supported, in part, by the National Health & Medical Research Council of Australia (program Grant #487922 (R.J.S), the Australian government Endeavour International Postgraduate Research Scholarship and the University of Melbourne International Research Scholarship (Y-S.C). Analysis of proteomic data described in this work was supported using the Australian Proteomics Computational Facility funded by the National Health & Medical Research Council of Australia Grant #381413. This work was supported by funds from the Operational Infrastructure Support Program provided by the Victorian Government, Australia. We acknowledge the Australian Cancer Research Foundation for providing funds to purchase the Orbitrap (TM) mass spectrometer. The authors thank Dr. Lifeng Peng and Prof. Bill Jordan for the microsome samples, and Robert Goode for access to the GRAVY software.