Mass spectrometric studies of human orosomucoid

Orosomucoid (OMD) was purified from pooled human plasma and characterized by mass spectrometry. OMD was subfractionated into two major gene products by immobilized metal affinity chromatography (IMAC) and they were also characterized by mass spectrometry and the results of the three preparations were compared. Matrix-assisted laser desorption ionization mass spectrometry (MALDI-TOF/MS) provided an accurate representation of the average molecular weight for OMD and was in agreement with the average molecular weight of the gene products. OMD and the individual gene products were subjected to enzymatic digestion followed by MALDI-TOF/MS, high performance liquid chromatography (HPLC) with on-line electrospray ionization mass spectrometry (ESI/MS) and fraction collection for off-line MALDI-TOF/MS. The on-line analysis provided detailed peptide maps for OMD and the individual gene products confirming the peptide sequence as determined from the OMD genes. The ESI/MS experiment was optimized to detect glycopeptide specific fragment ions and provided a means to locate glycopeptide containing HPLC fractions for further structural analysis. Based on the observation of an 803 m/z ion in glycopeptide spectra, a tandem mass spectrometry experiment was designed to detect the presence of the sialyl Lewis...