Journal article
Expression and purification of the human homeodomain oncoprotein HOX11
Protein Expression and Purification, Vol.25(2), pp.313-318
2002
Abstract
HOX11 is a transcription factor belonging to the homeodomain family that is essential for spleen development during embryogenesis. It is also tumorigenic, being associated with T-cell acute lymphoblastic leukemia in children. In order to understand the functional role of HOX11 in both normal development and malignancy, protein-DNA and protein-protein interaction studies involving this factor are required. Such investigations would be facilitated by the availability of significant amounts of purified HOX11 protein. However, expression of full-length HOX11 in bacteria has been reported to be problematic owing to fusion protein instability. Here, we report the purification of human HOX11 expressed in Escherichia coli as a soluble and functional glutathione S-transferase (GST) fusion protein. In addition, a mutant version of HOX11 was produced (HOX11ΔH3) which lacked the DNA-recognition helix (helix 3) of the homeodomain. Through a single purification procedure using glutathione-Sepharose, 2 mg of the recombinant proteins were obtained per liter of bacterial culture. Notably, recombinant GST-HOX11 fusion proteins had a markedly higher stability when purified at low temperature (4°C). Purification to near-homogeneity was achieved as judged by SDS-PAGE and the purified proteins were recognized by anti-HOX11 antibodies. The biological activity of the recombinant protein was verified by the specific binding of GST-HOX11, but not GST-HOX11ΔH3, to DNA containing consensus HOX11 recognition sites.
Details
- Title
- Expression and purification of the human homeodomain oncoprotein HOX11
- Authors/Creators
- M. Heidari (Author/Creator) - Murdoch UniversityK.L. Rice (Author/Creator) - Murdoch UniversityU.R. Kees (Author/Creator) - The Kids Research Institute AustraliaW.K. Greene (Author/Creator) - Murdoch University
- Publication Details
- Protein Expression and Purification, Vol.25(2), pp.313-318
- Publisher
- Academic Press
- Identifiers
- 991005540731007891
- Copyright
- © 2002 Elsevier Science (USA).
- Murdoch Affiliation
- School of Veterinary and Biomedical Sciences
- Language
- English
- Resource Type
- Journal article
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