Resumo

The members of the Paracoccidioides complex are the etiological agents of the systemic mycosis paracoccidioidomycosis. During the infection process the fungus requires several virulence factors such as adhesion, colonization of host cells and the ability to acquire nutrients. The plasma membrane is involved in different processes during the establishment of infection, such as transport of nutrients, the homeostatic regulation of the cell and in signaling cascades. It consists of a lipid bilayer with proteins associated. The membrane proteins can be classified into three categories depending on their association with the plasma membrane: integral membrane proteins, peripheral proteins and proteins attached to the membrane by a GPI anchor. NanoUPLC-MS^E technique was employed in order to identify membrane proteins of Paracoccidioides (Pb01) yeast cells grown in chemically defined media. The membrane protein extract was obtained after two-step of ultracentrifugation and subjected to tryptic digestion. After, the samples were analyzed by NanoUPLC-MS^E. Transmission Electron Microscopic (TEM) was performed in order to confirm the presence of membranes obtained by this protocol. In silico analyzes were performed to determine the location and possible association of the 167 identified proteins in the plasma membrane fraction. From the total, 48 proteins were predicted to be localized at the plasma membrane, inner or outer mitochondrial membrane and integral proteins, which comprehend one or more hydrophobic domains that cross the plasma membrane. Among the proteins classified as belonging to the membrane, 17 possesses transmembrane domains. Transport proteins SEC 61, ATPase plasma membrane and ATPase sodium transport have been identified as integral membrane proteins. The proteins hsp70, PCI domain containing and calnexin presented transmembrane domains and an N-terminal signal peptide. The knowledge of the constitution of the membrane is important to understand some processes developed by Paracoccidioides for its growth and survival within the host.