A simple method for recombinant protein purification using self-assembling peptide-tagged tobacco etch virus protease

Li, Guang‐Ya; Xiao, Zhenzhen; Lu, Huipeng; Li, Yangyang; Zhou, Xiaohui; Tan, Xiaoheng; Zhang, Xinyu; Xia, Xiaoli; Sun, Huaichang

doi:10.1016/j.pep.2016.08.013

Cited by 9 publications

(6 citation statements)

References 25 publications

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“…1b). This observation is in agreement with previous results for the expression of the natural DNA sequence of the bovine IFN- gene in E. coli in which Histagged rBoIFN- was purified by affinity chromatography from the soluble cell fraction [62]. The presence of the APPs in the recombinant protein caused a noticeable shift of the final products toward the insoluble cell fraction, as expected (Fig.…”

Section: Production Of Rboifn- In L Lactissupporting

confidence: 93%

Aggregation-prone peptides modulate activity of bovine interferon gamma released from naturally occurring protein nanoparticles

et al. 2020

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Section: Production Of Rboifn- In L Lactissupporting

confidence: 93%

Aggregation-prone peptides modulate activity of bovine interferon gamma released from naturally occurring protein nanoparticles

et al. 2020

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“…In case of C19 oxidized to the sulphenic acid, protein folding will be probably impaired, due to the TEVp 5M C19S produced as inclusion bodies. The insoluble TEVp construct retains the cleavage activity [39]. Further study will focus on construction of the cysteine-free TEVp variant aggregate for tag-removal.…”

Section: Discussionmentioning

confidence: 99%

Exploring the Mutations on Improving Oxidative Stability of Tobacco Etch Virus Protease for Tag-removal in Refolding of Two Disulfide-rich Proteins

Fan

Bayar

Ren

et al. 2021

Preprint

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Tobacco etch virus protease (TEVp) is a useful tool for removing fusion tag, but wild type TEVp shows less oxidative stability, which limits its application under the oxidized redox state to facilitate disulfide bonds formation for refolding disulfide-bonded proteins. Previously, we combined six mutations into the TEVp to generate the TEVp5M for obviously increasing the protein solubility and decreasing the auto-cleavage. In this work, we introduced and combined C19S, C110S and C130S mutations into the TEVp5M to generate seven variants, analyzed protein solubility and the cleavage activity of the constructs in each of three E. coli strains including BL21(DE3), BL21(DE3)pLys, and Rossetta(DE3), and those of the optimized soluble variants in the oxidative cytoplasm of Origami(DE3) under the same induction conditions. The results suggested that desirable protein solubility, cleavage activity and oxidative stability are not combined. Unlike that of the C19S, introduction of the C110S and/or C130S less affected protein solubility but increased tolerance to the oxidative redox state. Use of the TEVp5MC110S/C130S variant, the refolded disulfide-rich bovine enteropeptidase or maize peroxidase was released via cleaving the sequence between the target protein and the cellulose-binding module bound to regenerated amorphous cellulose.

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“…E. coli has served as a cell factory for recombinant protein expression for a long time and the cells expressing an enzyme can be used as biocatalysts [ 4 , 5 ], but overexpression of heterogeneous proteins can lead to the formation and accumulation of inactive IBs due to misfolded polypeptides [ 12 , 59 ]. Currently effective strategies have been developed to recover enzyme activity from aggregate particles [ 23 , 56 ]. Active IBs have shown unique advantages, such as easy separation, greater stability, and reusability, and offer robustness in applications [ 32 ].…”

Section: Discussionmentioning

confidence: 99%

Active tyrosine phenol-lyase aggregates induced by terminally attached functional peptides in Escherichia coli

Han

Zeng

Zhang

et al. 2020

Journal of Industrial Microbiology and Biotechnology

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The formation of inclusion bodies (IBs) without enzyme activity in bacterial research is generally undesirable. Researchers have attempted to recovery the enzyme activities of IBs, which are commonly known as active IBs. Tyrosine phenol-lyase (TPL) is an important enzyme that can convert pyruvate and phenol into 3,4-dihydroxyphenyl-l-alanine (L-DOPA) and IBs of TPL can commonly occur. To induce the correct folding and recover the enzyme activity of the IBs, peptides, such as ELK16, DKL6, L6KD, ELP10, ELP20, L6K2, EAK16, 18A, and GFIL16, were fused to the carboxyl terminus of TPL. The results showed that aggregate particles of TPL-DKL6, TPL-ELP10, TPL-EAK16, TPL-18A, and TPL-GFIL16 improved the enzyme activity by 40.9%, 50.7%, 48.9%, 86.6%, and 97.9%, respectively. The peptides TPL-DKL6, TPL-EAK16, TPL-18A, and TPL-GFIL16 displayed significantly improved thermostability compared with TPL. L-DOPA titer of TPL-ELP10, TPL-EAK16, TPL-18A, and TPL-GFIL16, with cells reaching 37.8 g/L, 53.8 g/L, 37.5 g/L, and 29.1 g/L, had an improvement of 111%, 201%, 109%, and 63%, respectively. A higher activity and L-DOPA titer of the TPL-EAK16 could be valuable for its industrial application to biosynthesize L-DOPA.

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A simple method for recombinant protein purification using self-assembling peptide-tagged tobacco etch virus protease

Cited by 9 publications

References 25 publications

Aggregation-prone peptides modulate activity of bovine interferon gamma released from naturally occurring protein nanoparticles

Aggregation-prone peptides modulate activity of bovine interferon gamma released from naturally occurring protein nanoparticles

Exploring the Mutations on Improving Oxidative Stability of Tobacco Etch Virus Protease for Tag-removal in Refolding of Two Disulfide-rich Proteins

Active tyrosine phenol-lyase aggregates induced by terminally attached functional peptides in Escherichia coli

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