ELECTROSPRAY-IONIZATION MASS-SPECTROSCOPY ON HYDROPHOBIC PEPTIDES ELECTROELUTED FROM SODIUM DODECYL-SULFATE POLYACRYLAMIDE-GEL ELECTROPHORESIS APPLICATION TO THE TOPOLOGY OF THE SARCOPLASMIC-RETICULUM CA2+ ATPASE

We describe a method to prepare proteins and pep-tides in a state suitable for exact determination of molecular mass by electrospray ionization mass spectrometry after sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and electroelution. The utility of the procedure, in conjunction with N-terminal sequencing, in defining the C-terminal end of the peptide fragments produced by proteolysis of sarcoplasmic reticulum Ca2+ ATPase with V8 is demonstrated. The application of mass spectrometry aids significantly the use of proteolytic enzymes for topological studies of membrane proteins, and SDS-PAGE is preferable to reverse-phase HPLC for separation of membraneous, hydrophobic peptides and proteins. © 1993 Academic Press, Inc.