Expression of recombinant antioxidative peptides from algae in pichia pastoris GS115 and escherichia coli MG1655

Abstract

The peptide extracted from algae protein waste hydrolysate with the 11 amino acid sequence VECYGPNRPQF, named as AW peptide in this research, was found to be a potential antioxidant. In this study, recombinant Pichia pastoris GS115 and Escherichia coli MG1655 were constructed to provide a high-level expression of this antioxidative peptide. For P. pastoris, the 234 base pair DNA fragment containing six copies of the target peptide linked by the codons of lysine was integrated to the chromosome. However, the recombinant peptide could not be obtained. It might be due to the intracellularly accumulation of target peptide which could not be extracted due to the lack of a tagged marker. The construction of recombinant E. coli for expressing the target peptide was done by cloning the same DNA fragment into pQE-30Xa expression vector before being transformed into E. coli MG1655. The verified strain, named as AW strain, showed the expression at transcriptional level detected by RT-PCR. Moreover, the target peptide was in the soluble protein fraction but the protein yield was still low. The target protein band was verified by immunoblotting with the detection of antiHis-HRP antibody. The crude fraction containing recombinant AW peptide elution from the Ni2+ affinity column. The DPPH and ABTS scavenging activities of the recombinant AW peptide elution were lower while the in vitro protective effect on DNA damage induced hydroxyl radicals was better than those of chemical synthesized one. To improve the recombinant AW peptide production, the optimization and purification, along with other antioxidant properties, should be further investigated.