COMBINATION OF HIGH-PERFORMANCE ANION-EXCHANGE CHROMATOGRAPHY AND ELECTROSPRAY MASS-SPECTROMETRY FOR ANALYSIS OF THE IN-VITRO O-GLYCOSYLATED MUCIN MOTIF PEPTIDE

Reversed-phase high-performance liquid chromatography (HPLC) and high- performance anion-exchange chromatography (HPAEC) with pulsed amperome tric detection were developed for the study of products obtained from the in vitro O-glycosylation of a mucin motif peptide, TTSAPTTS, the m ost representative sequence encoded by the human gene MUC(5C). After i ncubation of the peptide, which is rich in clustered hydroxyamino acid s, by both human colonic and gastric microsomal homogenates, the glyco sylated products were separated by HPLC and HPAEC and analysed by elec trospray mass spectrometry (ES-MS). The combination of HPAEC and ES-MS was the approach used for evaluating the differences between the poly peptide N-acetylgalactosaminyltransferase activity in different digest ive tissues.