Isolation, Purification and Characterization of Keratinolytic Proteinase from Microsporum canis

摘要

A keratinolytic proteinase secreted by Microsporum canis in a broth containing human hair was purified 134-fold from the culture filtrate by ion-exchange chromatography using DEAE-Sephacel, CM-Sephadex C-50, and by Sephadex G-75 gel filtration. The purified enzyme was electrophoretically homogeneous with a molecular weight of 33,000. The enzyme had an optimum pH of 8.0, and the activity was stable in the alkaline pH range. Enzyme activity increased with temperature up to 35 ℃ and was stable up to 45 ℃. The keratinolytic activity was not affected by the addition of nonionic detergents, was activated by Mg2+, but inhibited by Zn2+. The purified enzyme was used to obtain guinea pig antiserum. The antiserum tested by double diffusion against the purified enzyme showed a single line of precipitation and completely neutralized the proteinase activity. This study reaffirms that the proteinase from M. canis may be a biochemical mechanism for the invasion of keratinized tissue, and could possibly play a role in the hypersensitivity reactions arising from superficial infections of this fungus.