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Nature Protocols 4, 1023–1037 (1 June 2009) | doi:10.1038/nprot.2009.84
Coupling surface plasmon resonance to mass spectrometry to discover novel protein|[ndash]|protein interactions
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Abstract
The elucidation of protein–protein interaction networks is a crucial task in the postgenomic era. In this protocol, we describe our approach to discover protein–protein interactions using the surface plasmon resonance technique coupled to mass spectrometry (MS). A peptide or a protein is immobilized on a sensor chip and then exposed to brain extracts injected through the surface of the chip by a microfluidic system. The interactions between the immobilized ligand and the extracts can be monitored in real time. Proteins interacting with the peptide/protein are recovered, trypsinated and identified using MS. The data obtained are searched against a sequence database using the Mascot 2.1 software. Control experiments using blank sensor chips and/or randomized peptides are carried out to exclude nonspecific interactors. The protocol can be carried out in
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