at Google ScholarTitle data
Lauber, Thomas ; Marx, Ute C. ; Schulz, Axel ; Kreutzmann, Peter ; Rösch, Paul ; Hoffmann, Silke:
Accurate Disulfide Formation in Escherichia coli : Overexpression and Characterization of the First Domain (HF6478) of the Multiple Kazal-Type Inhibitor LEKTI.
In: Protein Expression and Purification.
Vol. 22
(2001)
Issue 1
.
- pp. 108-112.
ISSN 1046-5928
DOI: https://doi.org/10.1006/prep.2001.1415
Abstract in another language
The human hemofiltrate peptide HF6478, a putative serine proteinase inhibitor, which is part of the precursor protein LEKTI, was cloned, overexpressed, and purified. HF6478 contains two disulfide bridges with 1-4, 2-3 connectivity, sharing partial homology to Kazal-type domains and other serine proteinase inhibitors. It was expressed as thioredoxin (Trx) fusion protein, and disulfide formation occurred in the oxidative cytoplasm of Escherichia coli Origami (DE3) strain which carries a trxB(-)/gor522(-) double mutation. The soluble fusion protein was purified using metal-chelating affinity chromatography. Cleavage of the Trx fusion protein with factor Xa and subsequent purification yielded the final product in amounts sufficient for structural studies. Characterization of recombinant HF6478 was done by amino acid sequencing, mass spectrometry, capillary zone electrophoresis, and CD spectroscopy. Taking the blood filtrate peptide HF6478 as example, we present a strategy which should facilitate the expression of different extracellular proteins in the E. coli cytoplasm.
Further data
| Item Type: | Article in a journal |
|---|---|
| Refereed: | Yes |
| Institutions of the University: | Faculties > Faculty of Biology, Chemistry and Earth Sciences > Department of Chemistry > Former Professors > Chair Biopolymers - Univ.-Prof. Dr. Paul Rösch Faculties Faculties > Faculty of Biology, Chemistry and Earth Sciences Faculties > Faculty of Biology, Chemistry and Earth Sciences > Department of Chemistry Faculties > Faculty of Biology, Chemistry and Earth Sciences > Department of Chemistry > Former Professors Faculties > Faculty of Biology, Chemistry and Earth Sciences > Department of Chemistry > Chair Biochemistry with an Emphasis on Biophysical Chemistry |
| Result of work at the UBT: | Yes |
| DDC Subjects: | 500 Science > 540 Chemistry 500 Science > 570 Life sciences, biology |
| Date Deposited: | 25 Jan 2019 09:05 |
| Last Modified: | 01 Apr 2022 07:25 |
| URI: | https://eref.uni-bayreuth.de/id/eprint/47028 |