Binding Constant Determination Studies Utilizing Recombinant ΔCREB Protein

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Abstract
In this study, we report the binding constants (Kd) of the cAMP-responsive element binding protein (ΔCREB) for various cAMP-response element (CRE) motifs. We utilized purified recombinant ΔCREB protein in binding reactions with natural CRE motifs found in the promoter of two neuropeptide hormone genes and with several variant CRE motifs. The Kd of ΔCREB for the perfectly palindromic CRE, TGACGTCA, found within the somatostatin promoter is estimated to be 5.0 × 10−9 M. The Kd of ΔCREB for the variant CRE motif TG_CGTCA found within the enkephalin promoter is calculated to be in the 3 × 10−8 M. These studies provide an in vitro quantitative assessment of the binding affinity of ΔCREB for various CRE motifs.