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Improve Protein Analysis with the New, Mass Spectrometry-Compatible ProteasMAX™ Surfactant
Sergei Saveliev1, Daniel Simpson1, William Daily2, Carolyn Woodroofe2, Dieter Klaubert2, Grzegorz Sabat3, Robert Bulliet1, and Keith V. Wood1
1Promega Corporation, 2Promega Biosciences,
3Mass Spectrometry Facility, Biotechnology Center, University of Wisconsin-Madison.
Incomplete solubilization and digestion and poor peptide recovery are frequent limitations in protein sample preparation for mass spectrometry (MS) analysis. Additives such as SDS or urea can improve protein solubilization and denaturation, but they tend to have negative effects on digestion and MS analysis. Here we present a novel acid- and thermo-labile surfactant, ProteasMAX™ Surfactant, that improves solubilization, digestion and peptide recovery while avoiding negative effects observed with common solubilizing agents. The surfactant is designed to degrade over the course of a typical sample preparation protocol, generating zwitterionic and neutral species that do not interfere with MS.
The improved solubilization properties of the surfactant allowed us to solubilize hydrophobic membrane proteins at room temperature
within 1 hour. 2D LC-MS/MS analysis of mouse membrane proteome demonstrated a 70% increase in proteome coverage when the surfactant was added to the sample preparation protocol as a complementary solubilizing agent to urea. We observed up to a 30-fold increase in the rate of proteolysis in trypsin digestion of myoglobin in the presence of the surfactant. In-gel digestion showed up to 1.8-fold increase in protein coverage and up to 6.9-fold increase in MASCOT score. Long peptides in the range of 2,500–4,000 Da, which typically are poorly recovered following in-gel digestion, were present in quantities sufficient for MS/MS analysis. Experiments with model peptides and protein showed that the surfactant also minimizes the loss of peptides and proteins due to adsorption onto laboratory plasticware.
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