Abstract
BackgroundMore than a dozen of fungal immunomodulatory proteins (FIPs) have been identified to date, most of which are from Ganoderma species. However, little is known about the similarities and differences between different Ganoderma FIPs’ bioactivities. In the current study, two FIP genes termed FIP-gap1 and FIP-gap2 from G. applanatum, along with LZ-8 and FIP-gsi, another two representative Ganoderma FIP genes from G. lucidum and G. sinense were functionally expressed in Pichia. Subsequently, bioactivities of four recombinant Ganoderma FIPs were demonstrated and compared.ResultsAll the four Ganoderma FIP genes could be effectively expressed in P. pastoris GS115 at expression levels ranging from 197.5 to 264.3 mg L− 1 and simply purified by one step chromatography using HisTrap™ FF prepack columns. Amino acid sequence analysis showed that they all possessed the FIP conserved fragments. The homologies of different Ganoderma FIPs were from 72.6 to 86.4%. In vitro haemagglutination exhibited that FIP-gap1, FIP-gsi and LZ-8 could agglutinate human, sheep and mouse red blood cells but FIP-gap2 agglutinated none. Besides, the immunomodulation activities of these Ganoderma FIPs were as: rFIP-gap2 > rFIP-gap1 > rLZ-8 and rFIP-gsi in terms of proliferation stimulation and cytokine induction on murine splenocytes. Additionally, the cytotoxic activity of different FIPs was: rFIP-gap1 > rLZ-8 > rFIP-gsi > rFIP-gap2, examined by their inhibition of three human carcinomas A549, Hela and MCF-7.ConclusionsTaken together, four typical Ganoderma FIP genes could be functionally expressed in P. pastoris, which might supply as feasible efficient resources for further study and application. Both similarities and differences were indeed observed between Ganoderma FIPs in their amino acid sequences and bioactivities. Comprehensively, rFIP-gaps from G. applanatum proved to be more effective in immunomodulation and cytotoxic assays in vitro than rLZ-8 (G. lucidum) and rFIP-gsi (G. sinense).
Highlights
More than a dozen of fungal immunomodulatory proteins (FIPs) have been identified to date, most of which are from Ganoderma species
Nine of those FIPs are from Ganoderma species including LZ-8 (FIP-glu, G. lucidum), LZ-9 (G. lucidum), FIP-gts (G. tsugae), FIP-gja (G. japonicum), FIP-gsi (G. sinense), FIP-gmi (G. microsporum), FIP-gas (G. astum), FIP-gbo (G. boninense), FIP-gat (G. atrum), FIP-gap1 and FIP-gap2 [32–35]
Our results indicated that differences existed between different Ganoderma FIPs despite of some similarities in their protein sequences and amino acid compositions
Summary
More than a dozen of fungal immunomodulatory proteins (FIPs) have been identified to date, most of which are from Ganoderma species. More and more attention has been paid to bioactive proteins from Ganodermas in recent years [19–22] Of all those bioactive proteins, fungal immunomodulatory proteins (FIPs) are a novelly-identified protein family, which share some amino acid sequence similarity and immunological response action to immunoglobulins [23]. Ever since the first FIP, known as LZ-8 or FIP-glu, was isolated from the mycelia of G. lucidum by Kino et al in last century [24], more than fifteen FIPs have been identified from different fungi to date [25– 31] Nine of those FIPs are from Ganoderma species including LZ-8 (FIP-glu, G. lucidum), LZ-9 (G. lucidum), FIP-gts (G. tsugae), FIP-gja (G. japonicum), FIP-gsi (G. sinense), FIP-gmi (G. microsporum), FIP-gas (G. astum), FIP-gbo (G. boninense), FIP-gat (G. atrum), FIP-gap and FIP-gap (both from G. applanatum) [32–35]. FIP-gmi could ablate cancer stemness and cisplatin resistance in oral carcinomas stem cells through IL-6/Stat signaling inhibition [37]
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