1. A simple method for recombinant protein purification using self-assembling peptide-tagged tobacco etch virus protease.
- Author
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Li, Guang-Ya, Xiao, Zhen-Zhen, Lu, Hui-Peng, Li, Yang-Yang, Zhou, Xiao-Hui, Tan, Xiao, Zhang, Xin-Yu, Xia, Xiao-Li, and Sun, Huai-Chang
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RECOMBINANT proteins , *TOBACCO etch virus , *PROTEOLYTIC enzymes , *BIOTECHNOLOGY , *PROTEIN expression - Abstract
Recombinant protein purification remains to be a major challenge in biotechnology and medicine. In this paper we report a simple method for recombinant protein purification using self-assembling peptide-tagged tobacco etch virus protease (TEVp). After construction of an N -terminal ELK16 peptide fusion expression vector, we expressed ELK16-TEVp fusion protein in E. coli . SDS-PAGE analysis showed that ELK16-TEVp was expressed as active protein aggregates which could be purified to 91% purity with 92% recovery by centrifugation in the presence 0.5% Triton X-100. By using His-tagged bovine interferon-γ (His-BoIFN-γ) as the substrate, we demonstrated that EKL16-TEVp had a protease activity of 1.3 × 10 4 units/mg protein with almost 100% cleavage efficiency under the optimized conditions. More importantly, EKL16-TEVp could be removed from the cleavage reaction by single-step centrifugation. After removing the His-tag by nickel-conjugated agarose bead absorption, the recombinant BoIFN-γ (rBoIFN-γ) was purified to 98.3% purity with 63% recovery. The rBoIFN-γ had an antiviral activity of 1.6 × 10 3 units/mg protein against vesicular stomatitis virus. These data suggest that ELK16-TEVp may become a universal tool for recombinant protein purification. [ABSTRACT FROM AUTHOR]
- Published
- 2016
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