Abstract
Background/Aim: Identification and characterization of membrane proteins is a crucial challenge in proteomics research. Thus, we designed a novel method to prepare proteins possessing extensive hydrophobic stretches for mass spectrometry studies, without sacrificing other classes of proteins. Materials and Methods: This method uses sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) separation and relies solely on a matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) instrument, the most common and easiest to use mass spectrometer. Results: Using this analytical procedure, a significant number of hydrophobic peptides were recovered, with no reduction in overall sequence coverage and with a good identification of transmembrane proteins sequence. Applying this method to the systematic identification of proteins located in lipid rafts, up to 47% of identified proteins were obtained with an improvement of sequence coverage. Conclusion: The procedure presented here is suitable for both identifying purified hydrophobic proteins and systematically investigating hydrophobic protein mixtures. It can be easily applied even in non-dedicated laboratories, such as those mostly devoted to clinical chemistry.
Footnotes
- Received October 2, 2009.
- Revision received March 15, 2010.
- Accepted March 16, 2010.
- Copyright© 2010 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved