Functional expression of FIP-gts, a fungal immunomodulatory protein from Ganoderma tsugae in Sf21 insect cells.
The mushrooms of diverse Lingzhi species have been traditionally consumed as luxurious functional food supplements in Chinese society. FIP-gts, a fungal immunomodulatory protein found in Song-Shan Lingzhi (Ganodera tsugae) has been proposed to possess therapeutic effects on cancer and autoimmune diseases. To produce active FIP-gts for evaluation of oral administration, a recombinant FIP-gts (rFIP-gts) fused with a 6His-tag at its C-terminus was expressed in Sf21 insect cells by the baculovirus expression system. High yield (about 70%) and purity (about 90%) of rFIP-gts was obtained by one-step nickel-affinity chromatography. The correctness of the harvested rFIP-gts was verified by Western blot and MALDI-MS analyses. Optimal expression of rFIP-gts was observed when the Sf21 cells were infected with multiplicity of infection of 10 for 72 h, and the yield was up to 47.2 microg/3 x 10(6) infected cells. The immunomodulatory activity of the purified rFIP-gts was detected as the induction of interleukin 2 released from murine splenocytes. Compared with the rFIP-gts produced in Escherichia coli cells, the rFIP-gts produced in Sf21 cells possessed evidently higher specific immunomodulatory activity.
Jinn
Tzyy-Rong
TR
Graduate Institute of Biotechnology, National Chung Hsing University, Taichung, Taiwan.
Wu
Chi-Ming
CM
Tu
Wu-Chun
WC
Ko
Jiunn-Liang
JL
Tzen
Jason T C
JT
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D016207:Cytokines / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism*; D032902:Ganoderma / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics; D037102:Lectins / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013154:Spleen / Q000378:metabolism; D018411:Spodoptera
D016207:Cytokines; D005656:Fungal Proteins; D037102:Lectins; D011994:Recombinant Proteins