Abstract
An elastolytic proteinase was isolated from Aspergillus fumigatus by column chromatography using diethylaminoethyl (DEAE)-Sephacel, carboxymethyl (CM)-Sephadex C-50 and Mono-S. Homogeneity was confirmed by the formation of a single band after disk polyacrylamide gel electrophoresis (PAGE). This enzyme had a molecular weight of 32, 000 Da as determined by sodium dodecyl sulfate (SDS)-PAGE.
The enzyme activity was inhibited by leupeptin, diisopropyl fluorophosphate (DFP), phenylmethan-esulfonylfluoride (PMSF), α1-antitrypsin, α2-macroglobulin and ulinastatin. However, neither ethylene-diaminetetraacetic acid (EDTA) nor dithiothreitol (DTT) showed any effect on it. This enzyme contained 366 amino acid residues and exhibited an isoelectric point of 9.1. The Aα chain of human fibrinogen was cleaved first, followed later by the Bβ and γ chains.
Elastolytic proteinase from A. fumigatus possesses proteolytic activity as demmonstrated by the hydrolysis of Asn(3)-Gln(4), Gly(8)-Ser(9), Glu(13)-Ala(14), Tyr(16)-Leu(17), Gly(20)-Glu(21), Tyr(26)-Thr(27), and Lys(29)-Ala(30) bonds of oxidized insulin B chain.
