ArticleMicrobial cell factories2021
Expression of the human molecular chaperone domain Bri2 BRICHOS on a gram per liter scale with an E. coli fed-batch culture.
Article in Microbial cell factories, 2021. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 7 papers.
What it found
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Who cites it
7 citing papers in PubMed, 12 citations in OpenAlex.
- Increased cytoplasmic expression of PETase enzymes in E. coli.Microbial cell factories · 2024Article
- Review
- A new kid in the folding funnel: Molecular chaperone activities of the BRICHOS domain.Protein science : a publication of the Protein Society · 2023Review
- Short hydrophobic loop motifs in BRICHOS domains determine chaperone activity against amorphous protein aggregation but not against amyloid formation.Communications biology · 2023Article
- Amyloid Fibril Formation of Arctic Amyloid-β 1-42 Peptide is Efficiently Inhibited by the BRICHOS Domain.ACS chemical biology · 2022Article
- A spider silk-derived solubility domain inhibits nuclear and cytosolic protein aggregation in human cells.Communications biology · 2022Article
- The dimerization mechanism of the N-terminal domain of spider silk proteins is conserved despite extensive sequence divergence.The Journal of biological chemistry · 2022Article
Corrections and comments
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Authors and funding
6 authors at 1 institution in 1 country.
Funding
Abstract
backgroundThe human Bri2 BRICHOS domain inhibits amyloid formation and toxicity and could be used as a therapeutic agent against amyloid diseases. For translation into clinical use, large quantities of correctly folded recombinant human (rh) Bri2 BRICHOS are required. To increase the expression and solubility levels of rh Bri2 BRICHOS it was fused to NT*, a solubility tag derived from the N-terminal domain of a spider silk protein, which significantly increases expression levels and solubility of target proteins. To increase the expression levels even further and reach the g/L range, which is a prerequisite for an economical production on an industrial scale, we developed a fed-batch expression protocol for Escherichia coli.
resultsA fed-batch production method for NT*-Bri2 BRICHOS was set up and systematically optimized. This gradual improvement resulted in expression levels of up to 18.8 g/L. Following expression, NT*-Bri2 BRICHOS was purified by chromatographic methods to a final yield of up to 6.5 g/L. After removal of the NT*-tag and separation into different oligomeric species, activity assays verified that different assembly states of the fed-batch produced rh Bri2 BRICHOS have the same ability to inhibit fibrillar and non-fibrillar protein aggregation as the reference protein isolated from shake flask cultures.
conclusionsThe protocol developed in this work allows the production of large quantities of rh Bri2 BRICHOS using the solubility enhancing NT*-tag as a fusion partner, which is required to effectively conduct pre-clinical research.
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