Resumo

The cell wall proteins of pathogenic fungi are important in several processes, such as adhesion, stress response and host tissue damage. Once it operates at the interface between fungus and the host its compounds are putative target sites for new antifungal drugs. The present study aims to describe Paracoccidioides cell wall proteins by using proteomic approaches. The cell wall proteins were obtained from mycelium and yeast cells and fractioned according to the interactions that they establish with cell wall. The fraction 1 (F1) includes the non-covalent bound proteins and those linked to the cell wall through disulfide bonds, extracted with SDS and reducing agents. The fraction (F2) includes cell wall proteins linked to β-1,3-glucan via alkali-label bond and are obtained by NaOH extraction. The fraction 3 (F3) is composed by proteins that are GPI (Glycosilfosfatidinositol) anchored and other proteins bound to β-1,3-glucan, and are extracted with HF-piridin. The protein extracts were submitted to trypsin digestion and identified by nanoscale liquid chromatography coupled to mass spectrometry nanoUPLC-MS^E. We identified 110 proteins in the F1, 26 proteins in the F2 and 6 proteins in the F3 samples in yeast cells. Also, we identified in mycelium 10 proteins in the F1, 13 proteins in the F2 and 7 proteins in the F3 fraction. Among the identified proteins it was found a transglycosylase, homologous to Crh1p which is a GPI anchored protein, known to be involved in attaching chitin to β-glucan. Adhesins previously described in Paracoccidioides such as enolase, glyceraldehyde-3-phosphate, alcohol dehydrogenase, fructose-1,6-biphosphate aldose and some chaperones were also identified. Moreover, we identified formamidase that was previously described as localized in the fungus cell wall and may be involved in tissue damage besides contributing with antigenic properties. This study of cell wall proteins of Paracoccidioides is important to elucidate the cell wall structure and composition, as well as the mechanisms used by this pathogenic fungus to establish infection.