title: Assessment of the Fusion Tags on Increasing Soluble Production of the Active TEV Protease Variant and Other Target Proteins in E. coli.
abstract: In this study, five fusion tags affecting soluble production and cleavage activity of the tobacco etch virus (TEV) protease (TEVp) variant in Escherichia coli strains BL21 (DE3) and Rosetta™ (DE3) are investigated. Combination of the augmenting rare transfer RNAs (tRNAs) and the fused expressivity tag (N-terminal seven amino acid residues of E. coli translation initiation factor II) promotes the soluble TEVp partner expressed at relatively high level. Attachment of the maltose-binding protein (MBP) tag increases soluble expression of the protease released from the fusion protein in E. coli cells, but the incorporated TEVp recognition sequence slightly decreases expressivity of the fusion construct. Except for the green fluorescent protein, the attached expressivity tag shows less efficiency than the MBP tag in enhancing expression levels of the selected five target proteins in the Rosetta™ (DE3) cells under different induction conditions. Our results identified that high-level production of the functional target protein as the fusion partner in E. coli is combined with the intrinsic property of fusion tag, fusion protein stability, inherent folding of target protein, rare tRNA abundance, and the incorporated linker. Purified TEVp fusion constructs with the N-terminal expressivity tag, as well as the MBP partner, are the ideal alternatives for removing fusion tag.
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, Anhui, 230036, People's Republic of China.
authors.forename: Xuelian
authors.identifier: http://orcid.org/0000-0002-3843-0212
authors.initials: X
authors.lastname: Yu
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, Anhui, 230036, People's Republic of China.
authors.forename: Jiaqi
authors.initials: J
authors.lastname: Sun
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, Anhui, 230036, People's Republic of China.
authors.forename: Weiyu
authors.initials: W
authors.lastname: Wang
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, Anhui, 230036, People's Republic of China.
authors.forename: Li
authors.initials: L
authors.lastname: Jiang
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, Anhui, 230036, People's Republic of China.
authors.forename: Beijiu
authors.initials: B
authors.lastname: Cheng
authors.affiliation: Key Laboratory of Crop Biology of Anhui Province, Anhui Agricultural University, Hefei, Anhui, 230036, People's Republic of China. fanjun@ahau.edu.cn.
authors.forename: Jun
authors.initials: J
authors.lastname: Fan
mesh_terms: D010450:Endopeptidases / Q000096:biosynthesis / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D058619:Maltose-Binding Proteins / Q000096:biosynthesis / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D012995:Solubility
chemical_list: D058619:Maltose-Binding Proteins; D011993:Recombinant Fusion Proteins; D010450:Endopeptidases; C099488:TEV protease
keywords: Escherichia coli; Fusion tags; Soluble production; TEV protease; Target proteins