Purification and characterization of a novel immunomodulatory protein from the medicinal mushroom Trametes versicolor

Bioactive proteins represent an important group of functional agents in medicinal mushrooms. Trametes versicolor (L.) Lloyd is a mushroom frequently used in traditional Chinese medicine for its anti-tumor and immunomodulatory activities. A new immunomodulatory protein from T. versicolor, named TVC, was purified by ammonium sulfate precipitation, ion-exchange chromatography and gel filtration chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of the purified protein revealed a single band with a molecular weight of 15.0 kD. Native polyacrylamide gel analysis revealed a band at 30 kD, indicating that TVC exists in solution as a homodimer. Isoelectric focusing showed that TVC was an acidic protein with an isoelectric point of 4.0. TVC was found to lack carbohydrate modifications (based on periodic acid/Schiff staining) and it does not agglutinate mouse red blood cells, suggesting that TVC is not a lectin-like protein. Biological activity assays demonstrated that TVC can enhance the proliferation of splenocytes, while it has no stimulatory effects on CD4(+) and CD8(+) T cells. TVC markedly increases the proliferation of human peripheral blood lymphocytes in a dose-dependent manner and enhances the production of both nitric oxide and tumor necrosis factor-alpha by lipopolysaccharide-induced murine macrophages. The results indicate that TVC is an immunostimulant that can boost immune response. Comparison of the N-terminal amino acid residues and mass spectrometry results with the protein database revealed no homologous proteins.