IDENTIFICATION OF HUMAN MYOCARDIAL PROTEINS SEPARATED BY 2-DIMENSIONAL ELECTROPHORESIS USING AN EFFECTIVE SAMPLE PREPARATION FOR MASS-SPECTROMETRY

Peptide mass fingerprinting is a powerful tool for the identification of proteins separated by two-dimensional gel electrophoresis (2-DE). T he identification of in-gel digested proteins by peptide mass fingerpr inting was significantly improved in comparison to blot-digests by usi ng a peptide-collecting device. This device allows the effective purif ication and concentration of enzymatic digests of low-intensity spots without expensive equipment and is described in detail. Sensitivity in the fmol range was demonstrated by unequivocal identification of bovi ne serum albumin after sodium dodecyl sulfate - polyacrylamide gel ele ctrophoresis. Furthermore the high performance liquid chromatography p attern of in-gel digests indicated a 2- to 3-fold higher yield of the separated peptides. Therefore, a higher amount of the peptides was ava ilable to perform N-terminal sequencing. The identification of 16 prot eins from a high-resolution 2-DE gel map of human myocardium tissue ha s been achieved by means of this technique. Three of these proteins we re associated with changes in spot intensity with dilated cardiomyopat hy.