Abstract
A high molecular form of human kidney cultured urokinase (T HMW-UK) was highly purified by affinity chromatography on (Nα-(ε-aminocaproyl)-DL-homoarginine hexylester)-Sepharose, followed by isoelectric focusing and Sephadex G-100 gel filtration. The molecular weight (53, 400 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis), amino acid composition, and kinetic parameters for Glu-plasminogen activation (Km=0.92μM, kcat=28.7min-1) of the enzyme resembled those of the high molecular form of urinary urokinase (U HMW-UK). However, some differences were found as follows. 1) T HMW-UK was more resistant to reduction, and the molecule did not completely dissociate into two protein bands (heavy and light chains) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. 2) The isoelectric point of T HMW-UK (pI=9.3) by isoelectric focusing was more alkali than that of U HMW-UK (pI 8.9). 3) The specific activity of T HMW-UK (63, 500IU/mg protein) was approximately half of that of U HMW-UK (100, 000-120, 000IU/mg protein).
The results indicate certain difference in physicochemical properties between T HMW-UK and U HMW-UK, and that some T HMW-UK is composed of single polypeptide chain or that some differences exist in the sugar composition of T HMW-UK and U HMW-UK.
