XI International Meeting on Paracoccidioidomycosis

XI International Meeting on Paracoccidioidomycosis

Resume:23-1

Poster (Painel)

23-1	CHARACTERIZATION OF CARBONIC ANHYDRASES OF Paracoccidioides brasiliensis
Authors:	Mariana Vieira Tomazett (UFG - Univarsidade Federal de Goiás) ; Alexandre Melo Bailão (UFG - Univarsidade Federal de Goiás) ; Elisa Flávia Cardoso Bailão (UFG - Univarsidade Federal de Goiás) ; Fabiana Fonseca Zanoelo (UFG - Univarsidade Federal de Goiás) ; Clayton Luiz Borges (UFG - Univarsidade Federal de Goiás) ; Célia Maria de Almeida Soares (UFG - Univarsidade Federal de Goiás)

Abstract

Paracoccidioidomycosis (PCM) is the most important endemic deep mycosis in Latin America. The disease is caused by the dimorphic fungus Paracoccidioides brasiliensis (Pb). Infection is acquired by inhalation of the airborne conidia, derived from the mycelial form of the fungus, which once in the alveoli transform into the yeast infective form. Clinical forms of the disease range from asymptomatic pulmonary lesions to systemic generalized infections. Understanding of the complex interactions between the fungus and its host must include the identification of gene expression patterns during infection. Carbonic anhydrase (CA) belongs to the family of zinc metalloenzymes that catalyzes the reversible hydratation of carbon dioxide to bicarbonate. CAs are distributed among all domains of life and are currently divided into five different, evolutionary unrelated classes (&alpha, &beta, &gama, &delta and &zeta), which independently evolved similar catalytic mechanisms. Transcriptional studies have shown that carbonic anhydrase of P. brasiliensis is expressed in yeast cells recovered from liver of infected mice. In the present work we characterized the cDNAs encoding for four carbonic anhydrases of P.brasiliensis (PbCA1, PbCA2, PbCA3, PbCA4). The complete cDNAs present 934, 986, 921 and 1112 nucleotides, coding for proteins with 283, 148, 256, 301 amino acids respectively. The sequences of PbCAs were compared with sequences from other organisms in databases. The anhydrases PbCA1, PbCA2 and PbCA3 belong to the family of &beta-CAs and PbCA4 of the &alpha-CAs. Recombinant PbCAs were obtained in heterologous systems with 32 kDa, 16 kDa, 28 kDa and 33 kDa respectively. Mass spectrometry analysis confirmed the sequences of the produced proteins. The expression of PbCAs was evaluated by using real-time RT-PCR in mycelium, yeast cells and mycelium to yeast transition. The PbCAs was also evaluated in yeast cells exposed to 5% CO2. In the presence of CO2, PbCA1 and PbCA2 expression was reduced whereas PbCA4 expression was increased. PbCA1 expression was induced during the mycelium to yeast transition; PbCA2 and PbCA4 expression were induced in yeast cells. The expression of those proteins in P. brasiliensis suggests differential roles in the fungal physiology.

Keyword: Carbonic anhydrase, Paracoccidioides brasiliensis, CO2

਀�