Molecular cloning and recombinant expression of a gene encoding a fungal immunomodulatory protein from Ganoderma lucidum in Pichia pastoris

The fungal immunomodulatory proteins (FIPs) are a new protein family identified from several edible and medical mushrooms and play an important role in anti-tumor, anti-allergy and immunomodulating activities. A gene encoding the FIP was cloned from the mycelia of Changbai Lingzhi (Ganoderma lucidum) and recombinant expressed in the Pichia pastoris expression system. SDS-PAGE, amino acid composition and circular dichroism analyses of the recombinant FIP (reFIP) indicated that the gene was correctly and successfully expressed. In vitro assays of biological activities revealed that the reFIP exhibited similar immunomodulating capacities as native FIPs. The reFIP significantly stimulated the proliferation of mouse spleen lymphocytes and apparently enhanced the expression level of interleukin-2 released from the mouse splenocytes. In addition, anti-tumor activity assay showed that the reFIP could inhibit the proliferation of human leukemia-NB4 by inducing the cell apoptosis to a degree of about 32.4%. Taken together, the FIP gene from Changbai G. lucidum has been integrated into the yeast genome and expressed effectively at a high level (about 191.2 mg l(-1)). The reFIP possessed very similar biological activities to native FIPs, suggesting its potential application as a food supplement or immunomodulating agent in pharmaceuticals and even medical studies.