ArticlemAbs2014
Degradation of C-terminal tag sequences on domain antibodies purified from E. coli supernatant.
Article in mAbs, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 7 papers.
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Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
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Who cites it
7 citing papers in PubMed, 14 citations in OpenAlex.
- A Novel Anti-CD47 Nanobody Tetramer for Cancer Therapy.Antibodies (Basel, Switzerland) · 2024Article
- Functional integration of protein A binding ability to antibody fragments for convenient and tag-free purification.Bioengineered · 2023Article
- Site-specificEJNMMI radiopharmacy and chemistry · 2023Article
- Uncovering temporospatial sensitive TBI targeting strategies via in vivo phage display.Science advances · 2022Article
- Phosphate-regulated expression of the SARS-CoV-2 receptor-binding domain in the diatom Phaeodactylum tricornutum for pandemic diagnostics.Scientific reports · 2022Article
- TSGIT: An N- and C-terminal tandem tag system for purification of native and intein-mediated ligation-ready proteins.Protein science : a publication of the Protein Society · 2021Article
- Selection of cell-type specific antibodies on tissue-sections using phage display.Journal of cellular and molecular medicine · 2015Article
Corrections and comments
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Authors and funding
4 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Expression of recombinant proteins often takes advantage of peptide tags expressed in fusion to allow easy detection and purification of the expressed proteins. However, as the fusion peptides most often are flexible appendages at the N- or C-terminal, proteolytic cleavage may result in removal of the tag sequence. Here, we evaluated the functionality and stability of 14 different combinations of commonly used tags for purification and detection of recombinant antibody fragments. The tag sequences were inserted in fusion with the c-terminal end of a domain antibody based on the HEL4 scaffold in a phagemid vector. This particular antibody fragment was able to refold on the membrane after blotting, allowing us to detect c-terminal tag breakdown by use of protein A in combination with detection of the tags in the specific constructs. The degradation of the c-terminal tags suggested specific sites to be particularly prone to proteolytic cleavage, leaving some of the tag combinations partially or completely degraded. This specific work illustrates the importance of tag design with regard to recombinant antibody expression in E. coli, but also aids the more general understanding of protein expression.
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Registered trials
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