Details
| Original language | English |
|---|---|
| Pages (from-to) | 123-130 |
| Number of pages | 8 |
| Journal | Applied Microbiology and Biotechnology |
| Volume | 101 |
| Issue number | 1 |
| Early online date | 20 Aug 2016 |
| Publication status | Published - Jan 2017 |
Abstract
In this study, we present the development of a process for the purification of recombinant human bone morphogenetic protein-2 (rhBMP-2) using mixed-mode membrane chromatography. RhBMP-2 was produced as inclusion bodies in Escherichia coli. In vitro refolding using rapid dilution was carried out according to a previously established protocol. Different membrane chromatography phases were analyzed for their ability to purify BMP-2. A membrane phase with salt-tolerant properties resulting from mixed-mode ligand chemistry was able to selectively purify BMP-2 dimer from refolding mixtures. No further purification or polishing steps were necessary and high product purity was obtained. The produced BMP-2 exhibited a biological activity of 7.4 × 105 U/mg, comparable to commercial preparations. Mixed-mode membrane chromatography can be a valuable tool for the direct purification of proteins from solutions with high-conductivity, for example refolding buffers. In addition, in this particular case, it allowed us to circumvent the use of heparin-affinity chromatography, thus allowing the design of an animal-component-free process.
Keywords
- Animal-component-free, BMP-2, Membrane adsorbers, Mixed-mode chromatography
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Biotechnology
- Immunology and Microbiology(all)
- Applied Microbiology and Biotechnology
Cite this
- Standard
- Harvard
- Apa
- Vancouver
- BibTeX
- RIS
In: Applied Microbiology and Biotechnology, Vol. 101, No. 1, 01.2017, p. 123-130.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Purification of bone morphogenetic protein-2 from refolding mixtures using mixed-mode membrane chromatography
AU - Gieseler, Gesa
AU - Pepelanova, Iliyana
AU - Stuckenberg, Lena
AU - Villain, Louis
AU - Nölle, Volker
AU - Odenthal, Uwe
AU - Beutel, Sascha
AU - Rinas, Ursula
AU - Scheper, Thomas
N1 - Funding Information: This study was funded by the German Federal Ministry of Education and Research (Bundesministerium für Bildung und Forschung, BMBF), as part of the research program cluster Biokatalyse2021 and with grant no. p43.
PY - 2017/1
Y1 - 2017/1
N2 - In this study, we present the development of a process for the purification of recombinant human bone morphogenetic protein-2 (rhBMP-2) using mixed-mode membrane chromatography. RhBMP-2 was produced as inclusion bodies in Escherichia coli. In vitro refolding using rapid dilution was carried out according to a previously established protocol. Different membrane chromatography phases were analyzed for their ability to purify BMP-2. A membrane phase with salt-tolerant properties resulting from mixed-mode ligand chemistry was able to selectively purify BMP-2 dimer from refolding mixtures. No further purification or polishing steps were necessary and high product purity was obtained. The produced BMP-2 exhibited a biological activity of 7.4 × 105 U/mg, comparable to commercial preparations. Mixed-mode membrane chromatography can be a valuable tool for the direct purification of proteins from solutions with high-conductivity, for example refolding buffers. In addition, in this particular case, it allowed us to circumvent the use of heparin-affinity chromatography, thus allowing the design of an animal-component-free process.
AB - In this study, we present the development of a process for the purification of recombinant human bone morphogenetic protein-2 (rhBMP-2) using mixed-mode membrane chromatography. RhBMP-2 was produced as inclusion bodies in Escherichia coli. In vitro refolding using rapid dilution was carried out according to a previously established protocol. Different membrane chromatography phases were analyzed for their ability to purify BMP-2. A membrane phase with salt-tolerant properties resulting from mixed-mode ligand chemistry was able to selectively purify BMP-2 dimer from refolding mixtures. No further purification or polishing steps were necessary and high product purity was obtained. The produced BMP-2 exhibited a biological activity of 7.4 × 105 U/mg, comparable to commercial preparations. Mixed-mode membrane chromatography can be a valuable tool for the direct purification of proteins from solutions with high-conductivity, for example refolding buffers. In addition, in this particular case, it allowed us to circumvent the use of heparin-affinity chromatography, thus allowing the design of an animal-component-free process.
KW - Animal-component-free
KW - BMP-2
KW - Membrane adsorbers
KW - Mixed-mode chromatography
UR - http://www.scopus.com/inward/record.url?scp=84982306238&partnerID=8YFLogxK
U2 - 10.1007/s00253-016-7784-1
DO - 10.1007/s00253-016-7784-1
M3 - Article
C2 - 27542381
AN - SCOPUS:84982306238
VL - 101
SP - 123
EP - 130
JO - Applied Microbiology and Biotechnology
JF - Applied Microbiology and Biotechnology
SN - 0175-7598
IS - 1
ER -