Molecular cloning, codon-optimized gene expression, and bioactivity assessment of two novel fungal immunomodulatory proteins from Ganoderma applanatum in Pichia

Fungal immunomodulatory proteins (FIPs) have been identified from a series of fungi, especially in Ganoderma species. However, little is known about the FIPs from G. applanatum . In this study, two novel FIP genes, termed as FIP-gap1 and FIP-gap2 , were cloned from G. applanatum , characterized and...

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Published in:Applied microbiology and biotechnology 2018-07, Vol.102 (13), p.5483-5494
Main Authors: Zhou, Siya, Guan, Shixin, Duan, Zuowen, Han, Xiao, Zhang, Xin, Fan, Wenli, Li, Haoge, Chen, Lijing, Ma, Hui, Liu, Hangmei, Ruan, Yanye, Lin, Jingwei
Format: Article
Language:English
Subjects:
Amino acids
Ascomycota
Basidiomycota
Biocompatibility
Biological activity
Biomedical and Life Sciences
Biotechnological Products and Process Engineering
Biotechnology
Cancer
Cell proliferation
Cloning
Codons
Cytotoxicity
Erythrocytes
Feasibility studies
Fungi
Ganoderma
Gene expression
Genetic aspects
Homology
Immunomodulation
Immunomodulators
Interleukin 2
Life Sciences
Microbial Genetics and Genomics
Microbiological research
Microbiology
Optimization
Peptides
Physiological aspects
Pichia pastoris
Proteins
Sheep
Splenocytes
Toxicity
Yeast
γ-Interferon
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cited_by cdi_FETCH-LOGICAL-c510t-88c609c15b3b6d52efb2878336e7e2a632617f14048841d63639f53f2d42816e3
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creator Zhou, Siya
Guan, Shixin
Duan, Zuowen
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Zhang, Xin
Fan, Wenli
Li, Haoge
Chen, Lijing
Ma, Hui
Liu, Hangmei
Ruan, Yanye
Lin, Jingwei
description Fungal immunomodulatory proteins (FIPs) have been identified from a series of fungi, especially in Ganoderma species. However, little is known about the FIPs from G. applanatum . In this study, two novel FIP genes, termed as FIP-gap1 and FIP-gap2 , were cloned from G. applanatum , characterized and functionally expressed after codon optimization in Pichia pastoris GS115. Results showed that FIP-gap1 and FIP-gap2 comprised 342-bp encoding peptides of 113 amino acids, which shared a high homology with other Ganoderma FIPs. The yield of recombinant FIP-gap1 and FIP-gap2 increased significantly after codon optimization and reached 247.4 and 197.5 mg/L, respectively. Bioactivity assay in vitro revealed that both rFIP-gap1 and rFIP-gap2 could agglutinate mouse, sheep, and human red blood cells. Besides, rFIP-gap1 and rFIP-gap2 obviously stimulated the proliferation of mouse splenocytes and enhanced IL-2 and IFN-γ release. Cytotoxicity detection indicated that IC 50 of rFIP-gap1 towards A549 and HeLa cancer cells were 29.89 and 8.34 μg/mL, respectively, whereas IC 50 of rFIP-gap2 to the same cancer cells were 60.92 and 41.05 μg/mL, respectively. Taken together, novel FIP gaps were cloned and functionally expressed in P. pastoris , which can serve as feasible and stable resources of rFIP gaps for further studies and potential applications.
doi_str_mv 10.1007/s00253-018-9022-5
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However, little is known about the FIPs from G. applanatum . In this study, two novel FIP genes, termed as FIP-gap1 and FIP-gap2 , were cloned from G. applanatum , characterized and functionally expressed after codon optimization in Pichia pastoris GS115. Results showed that FIP-gap1 and FIP-gap2 comprised 342-bp encoding peptides of 113 amino acids, which shared a high homology with other Ganoderma FIPs. The yield of recombinant FIP-gap1 and FIP-gap2 increased significantly after codon optimization and reached 247.4 and 197.5 mg/L, respectively. Bioactivity assay in vitro revealed that both rFIP-gap1 and rFIP-gap2 could agglutinate mouse, sheep, and human red blood cells. Besides, rFIP-gap1 and rFIP-gap2 obviously stimulated the proliferation of mouse splenocytes and enhanced IL-2 and IFN-γ release. Cytotoxicity detection indicated that IC 50 of rFIP-gap1 towards A549 and HeLa cancer cells were 29.89 and 8.34 μg/mL, respectively, whereas IC 50 of rFIP-gap2 to the same cancer cells were 60.92 and 41.05 μg/mL, respectively. 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Cytotoxicity detection indicated that IC 50 of rFIP-gap1 towards A549 and HeLa cancer cells were 29.89 and 8.34 μg/mL, respectively, whereas IC 50 of rFIP-gap2 to the same cancer cells were 60.92 and 41.05 μg/mL, respectively. Taken together, novel FIP gaps were cloned and functionally expressed in P. pastoris , which can serve as feasible and stable resources of rFIP gaps for further studies and potential applications.</abstract><cop>Berlin/Heidelberg</cop><pub>Springer Berlin Heidelberg</pub><pmid>29705959</pmid><doi>10.1007/s00253-018-9022-5</doi><tpages>12</tpages></addata></record>
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subjects Amino acids
Ascomycota
Basidiomycota
Biocompatibility
Biological activity
Biomedical and Life Sciences
Biotechnological Products and Process Engineering
Biotechnology
Cancer
Cell proliferation
Cloning
Codons
Cytotoxicity
Erythrocytes
Feasibility studies
Fungi
Ganoderma
Gene expression
Genetic aspects
Homology
Immunomodulation
Immunomodulators
Interleukin 2
Life Sciences
Microbial Genetics and Genomics
Microbiological research
Microbiology
Optimization
Peptides
Physiological aspects
Pichia pastoris
Proteins
Sheep
Splenocytes
Toxicity
Yeast
γ-Interferon
title Molecular cloning, codon-optimized gene expression, and bioactivity assessment of two novel fungal immunomodulatory proteins from Ganoderma applanatum in Pichia
url http://sfxeu10.hosted.exlibrisgroup.com/loughborough?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-24T23%3A11%3A06IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-gale_proqu&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Molecular%20cloning,%20codon-optimized%20gene%20expression,%20and%20bioactivity%20assessment%20of%20two%20novel%20fungal%20immunomodulatory%20proteins%20from%20Ganoderma%20applanatum%20in%20Pichia&rft.jtitle=Applied%20microbiology%20and%20biotechnology&rft.au=Zhou,%20Siya&rft.date=2018-07-01&rft.volume=102&rft.issue=13&rft.spage=5483&rft.epage=5494&rft.pages=5483-5494&rft.issn=0175-7598&rft.eissn=1432-0614&rft_id=info:doi/10.1007/s00253-018-9022-5&rft_dat=%3Cgale_proqu%3EA542672453%3C/gale_proqu%3E%3Cgrp_id%3Ecdi_FETCH-LOGICAL-c510t-88c609c15b3b6d52efb2878336e7e2a632617f14048841d63639f53f2d42816e3%3C/grp_id%3E%3Coa%3E%3C/oa%3E%3Curl%3E%3C/url%3E&rft_id=info:oai/&rft_pqid=2031880343&rft_id=info:pmid/29705959&rft_galeid=A542672453&rfr_iscdi=true