Abstract:

A novel, simple and fast method based on a quadrupole ion trap

(QIT) and a time-of-flight mass spectrograph was developed for the

glycosylation site and glycan structure analysis of glycoprotein by

in-solution digestion and in-gel digestion which did not need to enrich

and label the glycopeptide. Horseradish peroxidase (HRP) and RNase B were

utilised to optimize the conditions of sample pre-processing. The effects

of the conditions were investigated separately and the optimal conditions

were: heat denaturation of RNase B, chemical denaturation of HRP, RNase B

digestion with endoproteinase Lys-C, HRP digestion with trypsin, 12-16 h

of digestion, 50% acetonitrile-5% trifluoroacetic acid of the extraction

solution, sandwich spotting method of the sample.

Key words: glycoprotein structure, conditions of optimization , mass spectrometric analysis, sample pre-preparation