Intermolecular interactions of the malate synthase of Paracoccidioides spp
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2013
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Background: The fungus Paracoccidioides spp is the agent of paracoccidioidomycosis (PCM), a pulmonary mycosis
acquired by the inhalation of fungal propagules. Paracoccidioides malate synthase (PbMLS) is important in the
infectious process of Paracoccidioides spp because the transcript is up-regulated during the transition from
mycelium to yeast and in yeast cells during phagocytosis by murine macrophages. In addition, PbMLS acts as an
adhesin in Paracoccidioides spp. The evidence for the multifunctionality of PbMLS indicates that it could interact
with other proteins from the fungus and host. The objective of this study was to identify and analyze proteins that
possibly bind to PbMLS (PbMLS-interacting proteins) because protein interactions are intrinsic to cell processes, and
it might be possible to infer the function of a protein through the identification of its ligands.
Results: The search for interactions was performed using an in vivo assay with a two-hybrid library constructed in
S. cerevisiae; the transcripts were sequenced and identified. In addition, an in vitro assay using pull-down GST
methodology with different protein extracts (yeast, mycelium, yeast-secreted proteins and macrophage) was performed,
and the resulting interactions were identified by mass spectrometry (MS). Some of the protein interactions were
confirmed by Far-Western blotting using specific antibodies, and the interaction of PbMLS with macrophages was
validated by indirect immunofluorescence and confocal microscopy. In silico analysis using molecular modeling,
dynamics and docking identified the amino acids that were involved in the interactions between PbMLS and
PbMLS-interacting proteins. Finally, the interactions were visualized graphically using Osprey software.
Conclusion: These observations indicate that PbMLS interacts with proteins that are in different functional categories,
such as cellular transport, protein biosynthesis, modification and degradation of proteins and signal transduction. These
data suggest that PbMLS could play different roles in the fungal cell.
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Paracoccidioides spp, Protein-protein interactions, Malate synthase
Citação
OLIVEIRA, Karine Martins de; SILVA Neto, Benedito Rodrigues da; PARENTE, Juliana Alves; SILVA, Roosevelt Alves da; QUINTINO, Guilherme Oliveira; VOLTAN, Aline Raque; MENDES-GIANNINI, Maria José Soares; SOARES, Célia Maria de Almeida; PEREIRA, Maristela. Intermolecular interactions of the malate synthase of Paracoccidioides spp. BMC Microbiology, London, v. 13, n. 107, p. 1-14, 2013.