Mechanisms of Nucleosome Engagement by Superfamily 2 ATPases

Abstract

The nucleosome acidic patch, a highly conserved surface composed of negatively charged residues on the H2A–H2B dimer, serves as a key docking site for a wide variety of chromatin regulators. Proteins that engage this patch often use basic “arginine anchor” motifs to form electrostatic contacts that stabilize their association with the nucleosome. Although several histone and DNA modifying proteins have been shown to use this interaction mode, the full diversity and mechanistic consequences of acidic patch recognition remain poorly understood. This dissertation explores the structural and functional roles of acidic patch binding across three studies, highlighting how this interaction modulates chromatin engagement and activity. In Chapter 2, I investigate how the human chromatin remodeler CHD1 uses a conserved “anchor element” to engage the nucleosomal acidic patch. Using cryo-electron microscopy (cryo-EM), I captured multiple conformational states of the CHD1-nucleosome complex, revealing a stepwise nucleosomal recruitment mechanism and regulatory pause states. These findings show how CHD1 couples recognition of the acidic patch and extranucleosomal DNA with directional DNA translocation, offering new insight into how remodelers achieve specificity and control during chromatin remodeling. Chapter 3 expands on the concept of acidic patch binding by introducing a structure-based in silico screening platform. Using AlphaFold-Multimer, we predicted interactions between the H2A-H2B dimer and over 7,000 human nuclear proteins, identifying 41 previously uncharacterized acidic patch interactors. This screen provides a scalable approach to mapping nucleosome interactions and establishes a new framework for identifying potential chromatin interactors. Finally, in Chapter 4, I focus on one of the high-confidence hits from our screen: the E3 ubiquitin ligase and SF2-type ATPase SHPRH. I visualize the SHPRH-nucleosome complex by cryo-EM and show that SHPRH binds the acidic patch using multiple arginine anchors. Interestingly, SHPRH adopts a unique conformation distinct from other ATPase-containing chromatin factors and interacts with additional nucleosomal features including the H4 N-terminal tail. We explore how these interactions coordinate SHPRH’s activity and regulate its ATP hydrolysis. Together, these studies highlight the acidic patch as a critical hub for chromatin regulation and demonstrate how a combination of structural biology, computational prediction, and functional analysis can uncover new mechanisms of genome regulation. The work presented here not only advances our understanding of chromatin remodeling mechanism but also provides new tools and concepts for investigating the interactions of chromatin-associated factors with nucleosomes.