CHARACTERIZATION OF PLASMIN DIGESTED PEPTIDE MAPS FOR RECOMBINANT HIGH-MOLECULAR-WEIGHT UROKINASE AND PROUROKINASE - ITS USE FOR THE ESTIMATION OF LOW-MOLECULAR-WEIGHT UROKINASE IN PRO.UK HMW.UK

The generation of active two chain urokinase from the proenzyme pro-ur okinase by catalytic amounts of plasmin is an important regulatory ste p in plasminogen activation. The plasminogen activator, urokinase, con verts plasminogen to plasmin which in turn dissolves fibrin blood clot s. We have studied the conversion of pro-urokinase (Pro.UK) to high mo lecular weight urokinase (HMW.UK) by plasmin. This paper describes a p eptide map that separates various plasmin cleavage products of Pro.UK/ HMW-UK. A partially purified sample of HMW.UK containing trace amounts of plasmin was incubated at room temperature for 12 days. The digeste d peptide mixture after reduction and S-carbamide methylation was sepa rated using reversed phase high performance liquid chromatography. The peptides were collected and analyzed for their N-terminal amino acid sequences, and in some cases by plasma desorption mass spectrometry (P DMS). The plasmin map so generated is useful in investigating the inte grity of any HMW.UK produced via plasmin cleavage step.