PROTEASE-IV, A UNIQUE EXTRACELLULAR PROTEASE AND VIRULENCE FACTOR FROM PSEUDOMONAS-AERUGINOSA

Comparisons of virulence between a Pseudomonas parent strain and an is ogenic mutant devoid of protease TV have demonstrated a significant ro le for this enzyme during infection. We have characterized purified Ps eudomonas aeruginosa protease TV in terms of its biochemical and enzym atic properties, and found it to be a unique extracellular protease. T he N-terminal decapeptide sequence of protease IV is not homologous wi th any published protein sequence. Protease IV has a molecular mass of 26 kDa, an isoelectric point of 8.70, and optimum enzymatic activity at pH 10.0 and 45 degrees C. Purified protease TV demonstrates activit y for the carboxyl side of lysine-containing peptides and can digest a number of biologically important proteins, including immunoglobulin, complement components, fibrinogen, and plasminogen. Protease IV is not inhibited by thiol-, carboxyl-, or metalloproteinase inhibitors. The total loss of enzyme activity in the presence of N-p-tosyl-L-chloro-me thyl ketone and the partial inhibition of enzyme activity by diisoprop yl fluorophosphate or phenylmethylsulfonyl fluoride imply that proteas e TV is a serine protease. Inhibition by dithiothreitol and beta-merca ptoethanol suggests that intramolecular disulfide bonds are essential for enzyme activity. The characteristics of this enzyme suggest that i nhibitors of serine proteases could be developed into a medication des igned to arrest tissue damage during Pseudomonas infection.