Recombinant Expression and Bioactivity Comparison of Four Typical Fungal Immunomodulatory Proteins from Three Main Ganoderma Species

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).Electronic supplementary materialThe online version of this article (10.1186/s12896-018-0488-0) contains supplementary material, which is available to authorized users.

Building a Versatile Protein Production Platform Using Engineered Trichoderma reesei

“…Through medium optimization, the production level of LZ8 in our Δ10LZ8 reached 711 mg/L, which exceeds the highest yield of LZ8 (350 mg/L) reported for Pichia pastoris. HSA has been expressed in various expression systems and the yield of HSA in P. pastoris is the highest so far .…”

“…Through medium optimization, the production level of LZ8 in our Δ10LZ8 reached 711 mg/L, which exceeds the highest yield of LZ8 (350 mg/L) reported for Pichia pastoris. 41 HSA has been expressed in various expression systems and the yield of HSA in P. pastoris is the highest so far. 42 Although we have increased production by deleting the transcription regulator of extracellular protease Pea1, the yield of HSA in our constructed chassis is still quite low owing to its quick degradation by the extracellular proteases of T. reesei.…”

In Silico functional and phylogenetic analyses of fungal immunomodulatory proteins of some edible mushrooms

“…This clade (I) also indicates that members of Ganoderma genus are relatively conservative during evolution. Among the members of the Ganodermataceae, both FIP-Gapp1 and Gapp2 occurred together (as showed by Qu et al 2018) while However, Fip-Gapp1 and Fip-Gapp2 formed an interesting lineage with FIP-Cru and shared their similarity it, a member of the family Gomphidiaceae, which is also evident from the BLAST-2Sequence (Additional file 2: Fig. S2) sharing a similarity of 98.23% with Fip-Gapp1.…”

“…They comprise of both classical and non-classical human leukocyte antigens (HLA – A, B, C, E, G and G) (Garcia-lora et al 2003 ). Similarly, FIPs are also involved in boosting immune responses and found to be associated with anti-inflammatory, anti-tumor effects and hemagglutination (Qu et al 2018 ; Li et al 2019 ; Chalamaiah et al 2018 ). Therefore, the current study will aid in improvising the immune system using the FIPs as model protein sources in humans against the invading pathogens.…”

“…The FIPs are small molecular weight proteins ranging from 12 to 13 kDa having around 110–114 amino acids. Asp and Val are found in abundance more than other amino acids and lack Cys, His, and Met residues (Qu et al 2018 ; Ejike et al 2020 ). Amino acid, Asp acts as a common site for carbohydrate attachment required for the formation of glycoproteins while Val is essential amino acid having stimulatory activity.…”