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Overexpression, purification, and biophysical characterization of the heterodimerization domain of the core-binding factor beta subunit.
- Source :
- Journal of Biological Chemistry; January 1998, Vol. 273 Issue: 4 p2480-2487, 8p
- Publication Year :
- 1998
-
Abstract
- Core-binding factors (CBF) are heteromeric transcription factors essential for several developmental processes, including hematopoiesis. CBFs contain a DNA-binding CBF alpha subunit and a non-DNA binding CBF beta subunit that increases the affinity of CBF alpha for DNA. We have developed a procedure for overexpressing and purifying full-length CBF beta as well as a truncated form containing the N-terminal 141 amino acids using a novel glutaredoxin fusion expression system. Substantial quantities of the CBF beta proteins can be produced in this manner allowing for their biophysical characterization. We show that the full-length and truncated forms of CBF beta bind to a CBF alpha DNA complex with very similar affinities. Sedimentation equilibrium measurements show these proteins to be monomeric. Circular dichroism spectroscopy demonstrates that CBF beta is a mixed alpha/beta protein and NMR spectroscopy shows that the truncated and full-length proteins are structurally similar and suitable for structure determination by NMR spectroscopy.
Details
- Language :
- English
- ISSN :
- 00219258 and 1083351X
- Volume :
- 273
- Issue :
- 4
- Database :
- Supplemental Index
- Journal :
- Journal of Biological Chemistry
- Publication Type :
- Periodical
- Accession number :
- ejs7217870