[Expression and activity analysis of interferonalpha-con and thymosin-alpha1].
This study aimed to obtain recombinant fusion protein of thymosin alphal(TM-alpha1) and consensus IFNalpha (IFNalpha-con) which have bath TM-alpha1 and IFNalpha-con activities. The DNA sequence for the fusion protein was cloned into expression vector of pET-22b (+) and expressed in BL21 (DE3)-Codon plus-RP-X. The expressed product (TM-alpha1-IFN-con) was soluble, and amounted to more than 20% in total proteins of E. coli. By precipitation of (NH4)2SO4, hydrophobic interaction chromatography (HIC, Phenyl Sepharose 6 Fast Flow), anion-exchange chromatography (Q Sepharose Fast Flow), cation-exchange chromatography (SP Sepharose Fast Flow) and gel filtration (Sephadex G-75), it was purified to more than 96% purity. The activity of fusion protein for antivirus was tested by cytopathic-effect inhibition assay and activity for promoting lymphocyte proliferation was tested by cell proliferative assay. The activity for antivirus was higher than commercial IFNalpha1b and IFNalpha2a and activity for promoting lymphocyte proliferation was similar to commercial TM-alpha1. The fusion protein had better effect for anti-HBV in vitro, its effect was stronger than combination of IFNalpha and TM-alpha1 and cell toxicity was less than combination of IFNalpha and TM-alpha1. The above results show that it has effect bath antivirus of IFNalpha and promoting lymphocyte proliferation of the soluble fusion protein expressed in E. coli.
Chongqing Medical University, Chongqing 400016, China. lxj6422@yahoo.com.cn
Xianjun
X
Liu
Fangxin
F
Liu
Bo
B
Li
Huiyun
H
Zhou
Qinqin
Q
Wang
D000595:Amino Acid Sequence; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007370:Interferon Type I / Q000096:biosynthesis* / Q000235:genetics; D016898:Interferon-alpha; D051379:Mice; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology*; D011994:Recombinant Proteins; D000077596:Thymalfasin; D013947:Thymosin / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000235:genetics
D000998:Antiviral Agents; D007370:Interferon Type I; D016898:Interferon-alpha; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C050739:interferon alfacon-1; D013947:Thymosin; D000077596:Thymalfasin