High Level Expression Of Recombinant Chicken IFN-α Gene And Antiviral Activity

IFNs belong to the large class of glycoproteins known as cytokines. They are named after their ability to"interfere"with viral replication within host cells. Since interferon was found by Isaacs and Lindenmann in 1957, researchers paid more and more attention to interferon's genetic structure, mechanism of action, genetic engineering. IFNs are typically divided into two IFN classes: Type I IFN and Type II IFN, lately discovered IFN -λwas considered as a novel type interferon and named as Type III interferon. IFNs belonging to all IFN classes are very important to fight against viral infections.IFN-αis a kind of Type I interferon, which received much more attention upon for its comprehensive antiviral activity. IFN-αhas a number of hypotype. Each hypotype contains 165~166 amino acid residue and has similar structure (homology>70 %). Typical IFN-αtertiary structure is composed of 5 alpha helix which is connected with one long ring structure and three short ring structure.Interferon's antivirus affect doesn't kill the virus directly but rather through cells themselves to fight against viral infections. Interferons with autocrine or paracrine secretion affect connect with special recepter on cell membrane surface. After connecting with specific accepter, Interferon initiats cascading signal amplification course. Signal will be transmitted into cell nucleus which regulate and control a series of gene express. Then it triggers a series of physiologic and biochemical responses. Antiviral protein inducted by Interferon mainly has PKR which could inactivate some enzymes required by ribosome synthesize to cause inhibition reproduction of virus; the primary function of 2', 5'-oligoadenylate synthetase (OAS) is activating endogenous RNase L which can degrade viral mRNA; adenosine deaminase (ADAR) can modify virus RNA's A to I which can prevent viral protein synthesize. Mx protein is a kind of GTP conjugated protein which can connect with viral nucleic acid and damage virus capsid protein. All these proteins display significant function during immune response induced by virus infection.Our country has a long history on poultry industry, poultry's variety resources, meat and egg products or units of product all at the first place in the world, so it is innegligible to prophylaxis and treat different kinds of diseases. Recombinate IFN-αhas a broad-spectrum and high performance antiviral activity. It has the advantages of no drug tolerance and medicine residual comparing with conventional beast use chemistry pharmacon, vaccine and other antivirus biological products. But recombination IFN-αalso presents some problems, for example, it has a low yield and difficulties in renaturation and purification which can lead to exorbitant production costs etc.In this study, the chicken IFN-αgene was synthesized using optimized codons based on online analysis tools and bio-software and cloned into pWL expression vector for expression in Escherichia coli. SDS-PAGE analysis showed that the expressed protein had a molecular weight of 19 ku and accounted for 28.3% of the total cellular protein. The expression product existed mainly in the form of inclusion body. Dissolved inclusion body proteins were denatured and renatured simultaneously to purify the chicken IFN-αby gel filtration using Sephadex G50. The purity of the renatured ChIFN-αexceed 98% and the yield reached 47.1mg/L (34.14%). The synthetical ChIFN-αshowed strong antiviral activity of 1.12×108 U/mg on CEF/NDV culture.