Characterization of the tryptic map of recombinant DNA derived tissue plasminogen activator by high-performance...

A detailed tryptic map is presented for recombinant human tissue plasminogen activator (rt-PA). Electrospray ionization mass spectrometry is utilized as an on-line HPLC detector for tryptic mapping of this glycoprotein. The additional dimension provided by mass spectrometry gives considerably more detail about the complex tryptic map and significantly enhances the high-resolution chromatographic separation by distinguishing by mass any coeluting components. Through this improvement, the proline isomers of a tryptic peptide were observed eluting over a broad range of retention times. The glycopeptides of rt-PA are observed as well as any corresponding nonglycosylated peptides. In addition, the carbohydrate heterogeneity is readily observed, allowing analysis of the carbohydrate composition. The characteristic diagonal patterns formed by glycopeptides in a contour plot of the data allow rapid recognition of the glycopeptides.