The Codon Optimization and Fused Expression of LL-37 and IFN-α2a in Pichia pastoris GS115

Abstract

Abstract: To express the fussed protein of human LL-37 and IFN-α2a in Pichia pastoris with codon optimization. Recording to the codon preference of P. pastoris, the codons of LL-37 and IFN-α2a were optimized. A flexible linker of GlyGlyGlyGlySer was placed between LL-37 and IFN-α2a. The newly scheming DNA sequence was synthesized chemically. P. pastoris GS115LI was constructed by integrating the new gene into GS115 with help of plasmid pPIC9K. With the scanning of geneticin concentration ladder, two strains, named as GS115LI1 and GS115LI2, were gotten with high copy number. With SDS-PAGE detection, anti-virus activity detection and antimicrobial activity detection of broth, the results proved GS115LI1 and GS115LI2 not only expressed the fused proteins, but also maintained the activities of LL-37 and IFN-α2a respectively. After fermentation and induction of recombinant, the productivity of fusion protein was 819.1 mg/L. With salting out, hydrophobic chromatography and ionic exchange chromatography, the purity of fusion protein was 97%, recovery rate was 46.2% and the potency of IFN was 2.6×10⁸ IU/mg. After codon optimization, the fused protein of human LL-37 and IFN-α2a was expressed effectively in P. pastroris with antimicrobial activity of LL-37 and antifungus activity of IFN-α2a.

Key words: IFN-α2a , LL-37 , Fused protein, Expression

Zhang Mingjie. The Codon Optimization and Fused Expression of LL-37 and IFN-α2a in Pichia pastoris GS115[J]. Biotechnology Bulletin, 2014, 0(11): 206-213.

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