Multienzymatic strategy for increased sequence coverage of membrane proteins using optimized nLC-MALDI-MS/MS

摘要

The applicability of less specific proteases to the identification of membrane proteins has been demonstrated. In comparison with the standard protease trypsin, these proteases are able to cleave also in hydrophobic transmembrane regions. However, the resulting peptides are often hard to analyze using MALDI as ionization technique. The high number of theoretically possible peptides produced by unspecific cleavages is additionally complicating MASCOT MS/MS searches. In this work we present the optimized nLC-MALDI-MS/MS analysis of yeast plasma membrane digests using chymotrypsin, elastase, pepsin and trypsin and additional TMTzero-labeling of peptides. The impact of combining the information of all digests and the improved detection and fragmentation of hydrophobic peptides after labeling was investigated.