ArticleProtein expression and purification2014
Self-processing of a barley subtilase expressed in E. coli.
Article in Protein expression and purification, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 6 papers.
What it found
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The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
The trial behind it
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
6 citing papers in PubMed.
- Pectin under stress: Pectin methylesterases at the core of cell wall remodeling, integrity sensing, and signaling in plants.Plant communications · 2026Review
- Pull the fuzes: Processing protein precursors to generate apoplastic danger signals for triggering plant immunity.Plant communications · 2024Review
- The fungal subtilase AsES elicits a PTI-like defence response in Arabidopsis thaliana plants independently of its enzymatic activity.Molecular plant pathology · 2020Article
- Tobacco phytaspase: Successful expression in a heterologous system.Bioengineered · 2017Article
- Isolation and Characterization of a Thionin Proprotein-processing Enzyme from Barley.The Journal of biological chemistry · 2015Article
- A simple and efficient seamless DNA cloning method using SLiCE from Escherichia coli laboratory strains and its application to SLiP site-directed mutagenesis.BMC biotechnology · 2015Article
Corrections and comments
PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.
Authors and funding
4 authors.
Funding
Abstract
The barley protease BAJ93208 belongs to the subtilase family of serine proteases. We have expressed BAJ93208 in the cytoplasm of the Escherichiacoli strain SHuffle C3030 using a rhamnose-inducible promoter. The expression construct included a (His)6-tag at the N-terminus and a strep-tag at the C-terminus. Western blot analysis revealed that the protein was processed at the N- and C-terminus. To exclude that this processing was due to contaminating E. coli proteases, a mutated BAJ93208 protease was constructed. This inactive mutant was not processed, demonstrating that the processing was an autocatalytic process. To define the exact cleavage sites mass spectrometry was used which detected four differently processed versions of the protease. At the N-terminus, the self-processing removed the internal inhibitor and an additional 19 amino acids. At the C-terminus there was a cleavage site after Ala(765) which also removed the strep-tag. This explained the inability to detect the purified (His)6-BAJ93208-strep protease with an anti-strep-tag antibody. Finally, an additional alanine was removed either at the N-terminus (Ala(119)) or at the C-terminus (Ala(764)).
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.