NADH/NAD<sup>+</sup> binding and linked tetrameric assembly of the oncogenic transcription factors CtBP1 and CtBP2
Heidi Erlandsen, Anne M. Jecrois, Jeffry C. Nichols, James L. Cole, William E. Royer
Abstract
The activation of oncogenic C‐terminal binding Protein (CtBP) transcriptional activity is coupled with NAD(H) binding and homo‐oligomeric assembly, although the level of CtBP assembly and nucleotide binding affinity continues to be debated. Here, we apply biophysical techniques to address these fundamental issues for CtBP1 and CtBP2. Our ultracentrifugation results unambiguously demonstrate that CtBP assembles into tetramers in the presence of saturating NAD + or NADH with tetramer to dimer dissociation constants about 100 n m . Isothermal titration calorimetry measurements of NAD(H) binding to CtBP show dissociation constants between 30 and 500 n m , depending on the nucleotide and paralog. Given cellular levels of NAD + , CtBP is likely to be fully saturated with NAD under physiological concentrations suggesting that CtBP is unable to act as a sensor for NADH levels.