title: Distinct chitinases are expressed during various growth phases of the human pathogen Paracoccidioides brasiliensis.
abstract: The aim of this work was the partial purification and subsequent evaluation of chitinase expression during the various growth phases of Paracoccidioides brasiliensis. Initially, PbCTS1r was expressed as a recombinant protein and displayed enzymatic activity against 4-MU-[N-acetylglucosamine (GlcNAc)]3 and 4-MU-(GlcNAc)2. Two proteins, 45 kDa and 39 kDa in size, were partially purified from P. brasiliensis yeast crude extract using cation-exchange chromatography coupled with HPLC and were characterised as PbCTS1 and PbCTS2, respectively. Anti-PbCTS1r antibody recognised two proteins in the crude extracts of yeast and the transitional stage between mycelial and yeast phases. In crude extracts of mycelium, only the 45 kDa protein was detected. However, quantitative real-time polymerase chain reaction led to the detection of small quantities of Pbcts2 transcript in the mycelial phase. In the yeast cell wall extract, only the 39 kDa protein was detected. Moreover, both proteins were secreted by the yeast parasitic phase, suggesting that these proteins participate in the modulation of the fungal environment. Phylogenetic analysis of the predicted PbCTS1 and PbCTS2 proteins indicated that they code for distinct chitinases in P. brasiliensis. During evolution, P. brasiliensis could have acquired the paralogues Pbcts1 and Pbcts2 for growth and survival in diverse environments in both saprophytic and parasitic phases.
authors.affiliation: Laboratório de Biologia Molecular, Instituto de Ciências Biológicas, Universidade Federal de Goiás, Goiânia, GO, Brasil.
authors.forename: Lidiane Aparecida da Penha
authors.initials: LA
authors.lastname: Santana
authors.forename: Marilene Henning
authors.initials: MH
authors.lastname: Vainstein
authors.forename: Patrícia Kott
authors.initials: PK
authors.lastname: Tomazett
authors.forename: Ludier Kesser
authors.initials: LK
authors.lastname: Santos-Silva
authors.forename: Alfredo Miranda
authors.initials: AM
authors.lastname: Góes
authors.forename: Augusto
authors.initials: A
authors.lastname: Schrank
authors.forename: Célia Maria de Almeida
authors.initials: CM
authors.lastname: Soares
authors.forename: Maristela
authors.initials: M
authors.lastname: Pereira
mesh_terms: D002688:Chitinases / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D018076:DNA, Complementary / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D025282:Mycelium / Q000201:enzymology* / Q000254:growth & development; D010228:Paracoccidioides / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D010802:Phylogeny; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D018076:DNA, Complementary; D004271:DNA, Fungal; D002688:Chitinases