Purification and characterization of a secretory serine proteinase of Acanthamoeba healyi isolated from GAE

We purified and characterized a serine proteinase secreted by Acanthamoeba healyi to evaluate it as a possible virulence factor in the pathogenesis of granulomatous amoebic encephalitis (GAE). Ammonium sulfate precipitated cu lture supernatant of A. healyi OC-3A strain was purified by chromatography on CM-Sepharose, Sephacryl-S200, and Q-2 anion-exchange columns. The purifi ed 33-kDa enzyme had a pH optimum of 8.0 and a temperature optimum of 40 C. Phenylmethyl-sulfonylfluoride and diisopropyl fluorophosphate, serine prot einase inhibitors, diminished activity of the enzyme to near zero. In addit ion to types I and IV collagen and fibronectin, the main components of the extracellular matrix, other proteins such as fibrinogen, IgG, IgA, albumin, and hemoglobin were also degraded by the enzyme. The broad substrate speci ficity of this secreted serine proteinase suggests that it may play an impo rtant role in pathogenesis of GAE by A. healyi.