Session: Session 1

Preliminary study on multi-digestion approaches for enhancing the identification of membrane and plasma membrane proteins by mass spectrometry

The detection of membrane proteins is of importance to (i) determine some surface biomarkers, (ii) the expression pattern of cell specific plasma membrane, and (iii) identify potential candidates for targeted pharmacological approaches. Brain microvessel endothelial cells (ECs) holding the blood-brain barrier properties are an ideal experimental model, given the difficulty of transferring circulating compounds into the cerebral compartment, and the need to adapt innovative and specific approaches to deliver pharmacological compounds to the brain. The aim of this study is to optimize the identification of membrane proteins and plasma membrane proteins in particular through a multi-enzymatic approach during the digestion step of mass spectrometry (MS) analysis. Our preliminary results led on CD34+ ECs and through label-free MS analysis reveal a differential pattern of proteins after digestion with trypsin, trypsin + endolysin C, chymotrypsin and the combination of chymotrypsin, trypsin and endolysin C in favor of a better identification rate for trypsin + endolysin C. This initial purpose will open to some other approaches focused on the labeling of the cell plasma membrane to better sort-out the surface proteome of ECs.