The α and β Subunits of the GA-binding Protein Form a Stable Heterodimer in Solution

• Published in: The Journal of biological chemistry 2000-03, Vol.275 (11), p.7749
• Main Authors: Yurii Chinenov, Michael Henzl, Mark E. Martin
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.275.11.7749

We have studied the assembly of GA-binding protein (GABP) in solution and established the role of DNA in the assembly of the transcriptionally active GABPα 2 β 2 heterotetrameric complex. GABP binds DNA containing a single PEA3/Ets-binding site (PEA3/EBS) exclusively as the αβ heterodimer complex, but readily binds as the GABPα 2 β 2 heterotetramer complex on DNA containing two PEA3/EBSs. Positioning of the PEA3/EBSs on the same face of the DNA helix stabilizes heterotetramer complex binding. These observations suggest that GABPαβ heterodimers are the predominant molecular species in solution and that DNA containing two PEA3/EBSs promotes formation of the GABPα 2 β 2 heterotetrameric complex. We analyzed the assembly of GABPα 2 β 2 heteromeric complexes in solution by analytical ultracentrifugation. GABPα exists as a monomer in solution while GABPβ exists in a monomer-dimer equilibrium ( K d = 1.8 ± 0.27 μ m ). In equimolar mixtures of the two subunits, GABPα and GABPβ formed a stable heterodimer, with no heterotetramer complex detected. Thus, GABP exists in solution as the heterodimer previously shown to be a weak transcriptional activator. Assembly of the transcriptionally active GABPα 2 β 2 heterotetramer complex requires the presence of specific DNA containing at least two PEA3/EBSs.