Urokinase-related proteins in human urine. Isolation and characterization of single-chain urokinase (pro-urokinase) and urokinase-inhibitor complex.

Abstract

Urokinase-related proteins were purified from 60liter batches of human urine collected into the protease inhibitor aprotinin to prevent proteolytic degradation. Three homogeneous species were obtained by chromatography on zinc chelate-Sepharose, SP-Sephadex C-50, Sephadex G-100, benzamidine-Sepharose, and immunoad~rption on a murine anti-human urokinase monoclonal antibody. One urokinase-related protein with M, 95,000 representing a complex of two-chain urokinase with an inhibitor accounts for about 70% of the total urokinase-related antigen in urine. Nucleophilic agents dissociate the complex into active twochain urokinase and a protein with M, 45,000-50,O~~ which is immunologically distinct from urokinase. Approximately 25% of the urinary urokinase-related antigen represents a single-chain molecule with Mr 54,000. This highly purified single-chain molecule was obtained with a yield of 5 gglliter of urine. Only trace amounts (<5%) of the uroklnase-related antigen were recovered as free two-chain urokinase.