Analysis of protein-protein interactions in human plasma using non-denaturing two-dimensional electrophoresis and mass spectrometry

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Protein-protein interactions between human plasma proteins have been detected by micro two-dimensional electrophoresis (2-DE), comparing the patterns obtained under non-denaturing conditions and those obtained under partially denaturing conditions. The protein spots on the 2-DE gels, 157 spots on a non-denaturing micro 2-DE gel and 175 spots on a partially denaturing micro 2-DE gel, were subjected to structural analysis by mass spectrometry. For mass spectrometric analysis, matrix assisted laser desorption-time of flight mass spectrometry (MALDI-TOF MS) and database search (peptide mass fingerprinting, PMF) were employed. Integrating the information obtained, it was ascertained that structural information on human plasma proteins, including their status of complex-formation, can be analyzed. Although for the moment most of the protein researchers are using 2-DE under denaturing conditions, present study revealed that 2-DE under non-denaturing conditions will become more important by the advance of mass spectrometric methods in protein analysis.