title: 1,10-phenanthroline inhibits the metallopeptidase secreted by Phialophora verrucosa and modulates its growth, morphology and differentiation.
abstract: Phialophora verrucosa is one of the etiologic agents of chromoblastomycosis, a fungal infection that affects cutaneous and subcutaneous tissues. This disease is chronic, recurrent and difficult to treat. Several studies have shown that secreted peptidases by fungi are associated with important pathophysiological processes. Herein, we have identified and partially characterized the peptidase activity secreted by P. verrucosa conidial cells. Using human serum albumin as substrate, the best hydrolysis profile was detected at extreme acidic pH (3.0) and at 37 °C. The enzymatic activity was completely blocked by classical metallopeptidase inhibitors/chelating agents as 1,10-phenanthroline and EGTA. Zinc ions stimulated the metallo-type peptidase activity in a dose-dependent manner. Several proteinaceous substrates were cleaved, in different extension, by the P. verrucosa metallopeptidase activity, including immunoglobulin G, fibrinogen, collagen types I and IV, fibronectin, laminin and keratin; however, mucin and hemoglobin were not susceptible to proteolysis. As metallopeptidases participate in different cellular metabolic pathways in fungal cells, we also tested the influence of 1,10-phenanthroline and EGTA on P. verrucosa development. Contrarily to EGTA, 1,10-phenanthroline inhibited the fungal viability (MIC 0.8 µg/ml), showing fungistatic effect, and induced profound morphological alterations as visualized by transmission electron microscopy. In addition, 1,10-phenanthroline arrested the filamentation process in P. verrucosa. Our results corroborate the supposition that metallopeptidase inhibitors/chelating agents have potential to control crucial biological events in fungal agents of chromoblastomycosis.
authors.affiliation: Laboratório de Taxonomia, Bioquímica e Bioprospecção de Fungos, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, Brazil.
authors.forename: Marcela Queiroz
authors.initials: MQ
authors.lastname: Granato
authors.forename: Priscila de Araújo
authors.initials: Pde A
authors.lastname: Massapust
authors.forename: Sonia
authors.initials: S
authors.lastname: Rozental
authors.forename: Celuta Sales
authors.initials: CS
authors.lastname: Alviano
authors.forename: André Luis Souza
authors.initials: AL
authors.lastname: dos Santos
authors.forename: Lucimar Ferreira
authors.initials: LF
authors.lastname: Kneipp
mesh_terms: D000935:Antifungal Agents / Q000494:pharmacology*; D005656:Fungal Proteins / Q000378:metabolism*; D006801:Humans; D045726:Metalloproteases / Q000378:metabolism*; D009181:Mycoses / Q000382:microbiology; D010618:Phenanthrolines / Q000494:pharmacology*; D010676:Phialophora / Q000187:drug effects* / Q000201:enzymology* / Q000254:growth & development; D000069284:Protein Translocation Systems / Q000378:metabolism; D021381:Protein Transport; D013172:Spores, Fungal / Q000187:drug effects / Q000201:enzymology / Q000254:growth & development*
chemical_list: D000935:Antifungal Agents; D005656:Fungal Proteins; D010618:Phenanthrolines; D000069284:Protein Translocation Systems; D045726:Metalloproteases; C025205:1,10-phenanthroline