Secreted proteases from pathogenic fungi.

Biodegradation of keratin waste: Theory and practical aspects

Dermatophytes are highly specialized pathogenic fungi that exclusively infect the stratum corneum, nails or hair, and it is evident that secreted proteolytic activity is important for their virulence. Endo- and exoproteases-secreted by dermatophytes are similar to those of species of the genus Aspergillus. However, in contrast to Aspergillus spp., dermatophyte-secreted endoproteases are multiple and are members of two large protein families, the subtilisins (serine proteases) and the fungalysins (metalloproteases). In addition, dermatophytes excrete sulphite as a reducing agent. In the presence of sulphite, disulphide bounds of the keratin substrate are directly cleaved to cysteine and S-sulphocysteine, and reduced proteins become accessible for further digestion by various endo- and exoproteases secreted by the fungi. Sulphitolysis is likely to be an essential step in the digestion of compact keratinized tissues which precedes the action of all proteases.

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Secreted proteases from dermatophytes.

Trichophyton gallinae could grow on agar medium containing chicken feathers as sole sources of carbon and nitrogen, but it could not utilize human or guinea pig hair. The fungus grows rapidly when feather keratin solubilized in dimethyl sulphoxide was included in the growth medium. Keratinase activity could be detected by release of amino acids from solubilized keratin and by creation of zones of lysis in gel diffusion assay. However, the activity was observed only in homogenates of T. gallinae mycelia: it was not possible to detect it in culture filtrates.

Intracellular keratinase of Trichophyton gallinae.

Secreted subtilisin gene family in Trichophyton rubrum.

An exocellular proteinase produced by Trichophyton rubrum in a glucose-peptone broth was purified from lyophilized and dialysed culture filtrate of the dermatophyte by Sephadex G-100 gel filtration and preparative polyacrylamide gel electrophoresis. The purified enzyme was a homogeneous protein of molecular weight 34 700 and it could hydrolyse azoalbumin, casein, bovine serum albumin, α-N-benzoyl-L-arginine ethyl ester and p-toluenesulfonyl-L-arginine methyl ester but not N-benzoyl-L-tyrosine ethyl ester, α-N-benzoyl-DL-arginine-p-nitroanilide and keratin. The enzyme showed an alkaline pH optimum and was not activated by divalent metal ions but inhibited strongly by phenylmethylsulfonylfluoride. Thus the enzyme was identified as an alkaline serine proteinase.

S.K. Das

Papers

Purification and partial characterization of an exocellular proteinase from Trichophyton rubrum