Henley, M. J. | Search bioRxiv

bioRxiv2021

A Dual-Site Inhibitor of CBP/p300 KIX is a Selective and Effective Modulator of Myb

The protein-protein interaction between the KIX motif of the transcriptional coactivator CBP/p300 and the transcriptional activator Myb is a high value target due to its established role in certain acute myeloid leukemias (AML) and potential contributions to other cancers. However, the CBP/p300 KIX domain has multiple binding sites, several structural homologues, many binding partners, and substantial conformational plasticity, making it challenging to specifically target using small molecule inhibitors. Here, we report a picomolar dual-site inhibitor (MybLL-tide) of the Myb-CBP/p300 KIX interaction. MybLL-tide has higher affinity for CBP/p300 KIX than any previously reported compounds while also possessing 16,000-fold selectivity for the CBP/p300 KIX domain over other coactivator domains. MybLL-tide blocks the association of CBP and p300 with Myb in the context of the proteome leading to inhibition of key Myb*KIX-dependent genes in AML cells. These results show that MybLL-tide is an effective, modifiable tool to selectively target the KIX domain and assess transcriptional effects in AML cells and potentially other cancers featuring aberrant Myb behavior. Additionally, the dual-site design has applicability to the other challenging coactivators that bear multiple binding surfaces

biochemistry↗

bioRxiv2020

Unexpected Specificity within Dynamic Transcriptional Protein-Protein Complexes

A key functional event in eukaryotic gene activation is the formation of dynamic protein-protein interaction networks between transcriptional activators and transcriptional coactivators. Seemingly incongruent with the tight regulation of transcription, many biochemical and biophysical studies suggest that activators use nonspecific hydrophobic and/or electrostatic interactions to bind to coactivators, with few if any specific contacts. Here a mechanistic dissection of a set of representative dynamic activator*coactivator complexes, comprised of the ETV/PEA3 family of activators and the coactivator Med25, reveals a different molecular recognition model. The data demonstrate that small sequence variations within an activator family significantly redistribute the conformational ensemble of the complex while not affecting overall affinity, and distal residues within the activator--not often considered as contributing to binding--play a key role in mediating conformational redistribution. The ETV/PEA3*Med25 ensembles are directed by specific contacts between the disordered activator and the Med25 interface, which is facilitated by structural shifts of the coactivator binding surface. Taken together, these data highlight the critical role coactivator plasticity plays in recognition of disordered activators, and indicates that molecular recognition models of disordered proteins must consider the ability of the binding partners to mediate specificity.

biochemistry↗