title: Identification of a metallopeptidase with TOP-like activity in Paracoccidioides brasiliensis, with increased expression in a virulent strain.
abstract: Paracoccidioidomycosis (PCM), caused by the pathogenic fungus Paracoccidioides brasiliensis, is a systemic mycosis with severe acute and chronic forms. The pathology of PCM is not completely understood, and the role of proteases in the infection is not clearly defined. In this report, we describe a metallopeptidase activity in P. brasiliensis total and cytosolic protein extracts similar to that of mammalian thimet oligopeptidase (TOP). The analogous enzyme was suggested by analysis of P. brasiliensis genome databank and by hydrolytic activity of the FRET peptide Abz-GFSPFRQ-EDDnp which was completely inhibited by o-phenanthrolin and significantly inhibited by the TOP inhibitor, JA-2. This activity was also partially inhibited by IgG purified from patients with PCM, but not from normal individuals. As shown by high-performance liquid chromatography (HPLC), the hydrolysis of bradykinin had the same pattern as that of mammalian TOP, and anti-mammalian TOP antibodies significantly inhibited fungal cytosolic peptidase activity. Moreover, anti-mammalian TOP antibodies recognized a component of 80 kDa on fungal cytosol. A P. brasiliensis virulent isolate showed higher gene expression and TOP-like peptidase activity than a non-virulent strain. The release of enzyme following fungal lysis would be consistent with host antibody production and may have a role in the pathogenesis, inflammation and further development of the mycosis.
authors: Unidade de Oncologia Experimental (UNONEX), Departamento de Microbiologia, Imunologia e Parasitologia, Universidade Federal de São Paulo-Escola Paulista de Medicina (UNIFESP-EPM), São Paulo, SP, Brazil.
authors: Ellen T
authors: ET
authors: Gravi
authors: Thaysa
authors: T
authors: Paschoalin
authors: Bianca R
authors: BR
authors: Dias
authors: Dayson F
authors: DF
authors: Moreira
authors: José E
authors: JE
authors: Belizario
authors: Vitor
authors: V
authors: Oliveira
authors: Adriana K
authors: AK
authors: Carmona
authors: Maria A
authors: MA
authors: Juliano
authors: Luiz R
authors: LR
authors: Travassos
authors: Elaine G
authors: EG
authors: Rodrigues
mesh_terms: D000818:Animals; D001920:Bradykinin / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004271:DNA, Fungal / Q000235:genetics; D004791:Enzyme Inhibitors / Q000378:metabolism; D020869:Gene Expression Profiling; D006801:Humans; D008168:Lung / Q000382:microbiology; D008297:Male; D045726:Metalloproteases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D010228:Paracoccidioides / Q000201:enzymology* / Q000302:isolation & purification / Q000472:pathogenicity*; D010229:Paracoccidioidomycosis / Q000382:microbiology; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid; D014774:Virulence
chemical_list: D004271:DNA, Fungal; D004791:Enzyme Inhibitors; D045726:Metalloproteases; D001920:Bradykinin