In vitro Synthesis of a Recombinant Fungal Immunomodulatory Protein from Lingzhi or Reishi Medicinal Mushroom, Ganoderma lucidum (W.Curt.:Fr.) P.Karst. (Aphyllophoromycetideae) and Analysis of Its Immunomodulatory Activity
In vitro Synthesis of a Recombinant Fungal Immunomodulatory Protein from Lingzhi or Reishi Medicinal Mushroom, Ganoderma lucidum (W.Curt.:Fr.) P.Karst. (Aphyllophoromycetideae) and Analysis of Its Immunomodulatory Activity
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灵芝或灵芝药用灵芝 (Ganoderma lucidum (W.Curt.:Fr.) P.Karst) 中重组真菌免疫调节蛋白的体外合成。
DOI：
10.1615/intjmedmushr.v12.i4.20
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发表时间：
2010-01-01
影响因子：
1.2
通讯作者：
Zhou, Xuan-wei
中科院分区：
文献类型：
--
作者：
Li, Qi-zhang;Wang, Xue-fei;Zhou, Xuan-wei
从药用蘑菇灵芝（灵芝或灵芝）中分离的FIP-glu（灵芝8或LZ-8）是第一个鉴定和表征的真菌免疫调节蛋白（FIP），其生物学功能已被广泛探索。基于从灵芝基因组DNA克隆的LZ-8基因序列，我们通过表达盒载体pQE-30表达FIP-glu。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）和基质辅助激光解吸/电离质谱（MALDI-MS）分析重组蛋白。最后，通过诱导小鼠脾细胞中细胞因子基因的表达来检查其生物活性。结果表明重组FIP-glu蛋白能够在大肠杆菌中成功表达。重组FIP-glu的产量占大肠杆菌总蛋白的51.1%，可溶性重组FIP-glu占大肠杆菌可溶性蛋白总量的55.4%。重组FIP-glu蛋白纯化后纯度达90%以上。 RT-PCR分析表明，重组FIP-glu可以增强小鼠脾细胞中白细胞介素（IL）-2、IL-3、IL-4、干扰素（IFN）-γ、肿瘤坏死因子（TNF）-α和IL-2受体（IL-2R）基因的转录。
The FIP-glu (Lingzhi-8 or LZ-8) isolated from medicinal mushroom Ganoderma lucidum (Lingzhi or Reishi) was the first identified and characterized fungal immunomodulatory protein (FIP), and its biological functions have been explored extensively. On the basis of cloned LZ-8 gene sequence from the genomic DNA of G. lucidum, we expressed FIP-glu by the expression cassette vector pQE-30. The recombinant protein was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Finally, its bioactivity was examined by inducing expression of cytokine genes in mouse spleen cells. The results showed that recombinant FIP-glu protein could be expressed in E. coli successfully. The yield of recombinant FIP-glu accounted for 51.1% of the total protein of E. coli, while soluble recombinant FIP-glu accounted for 55.4% of the total soluble protein of E. coli. The purity of recombinant FIP-glu protein was more than 90% after purification. Analysis of RT-PCR demonstrated that the recombinant FIP-glu could enhance the transcription of interleukin (IL)-2, IL-3, IL-4, interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, and IL-2 receptor (IL-2R) genes in mouse spleen cells.