WDR5 Binding to Histone Serotonylation Is Driven by an Edge-Face Aromatic Interaction with Unexpected Electrostatic Effects

Abstract

Histone serotonylation has emerged as a key post-translational modification. WDR5 preferentially binds to serotonylated histone 3 (H3), and this binding event has been associated with tumorigenesis. Herein, we utilize genetic code expansion, structure-activity relationship studies, and computation to study an edge-face aromatic interaction between WDR5 Phe149 and serotonin on H3 that is key to this protein-protein interaction. We find experimentally that this edge-face aromatic interaction is unaffected by modulating the electrostatics of the face component but is weakened by electron-withdrawing substituents on the edge component. Overall, these results elucidate that this interaction is governed by van der Waals forces as well as electrostatics of the edge ring, a result that clarifies discrepancies among previous theoretical models and model system studies of this interaction type. This is the first evaluation of the driving force of an edge-face aromatic interaction at a protein-protein interface and provides a key benchmark for the nature of these understudied interactions that are abundant in the proteome.

Figure 1.

Common geometries of aromatic interactions.

Figure 2.

Crystal structure of WDR5 binding H3Q5sero (PDB 7CFP) with (a) top-down view of whole protein and (b) molecular level interactions with serotonin and residue Phe149 of WDR5 engaging in an edge-face aromatic interaction, viewed from the side relative to (a).

Figure 3.

Bar graph showing energy decomposition analysis (EDA) of WDR5 Phe149 interacting with serotonin calculated at the M06–2X/6–311++G(d,p)/SMD(Water) level of theory.

Figure 4.

Structures, ESP values, and ESP maps of para-substituted Phe analogs used in this study. ESP maps were calculated in Spartan using the DFT ωB97X-D/6–31G(d) level of theory with an energetic range from −100 to +100 kcal/mol. Blue indicates positive ESP, green is neutral, and red indicates negative ESP. Ring center ESP values were previously reported. See Supporting Information for details on calculations of ortho edge H ESP values. All ESP values shown are in units of kcal/mol. A portion of this figure was adapted from reference . Copyright 2022 American Chemical Society.

Figure 5.

LFER plots evaluating the correlation between ΔG_binding to H3Q5sero and the varying ESP of Phe149 of WDR5.

Figure 6.

Structures, ESP values, and ESP maps of serotonin and its analogs used in this study. ESP maps were calculated in Spartan using the DFT ωB97X-D/6–31G(d) level of theory with an energetic range from −100 to +100 kcal/mol. Blue indicates positive ESP, green is neutral, and red indicates negative ESP. See Supporting Information for details on calculations of ESP values.

Figure 7.

LFER plot showing that ΔG_binding to WDR5 WT and WDR5 F149pClF is unaffected by changing in electrostatics of the serotonin moiety. Each data point is labeled with the identity of the substituent at the 5 position of tryptamine.