Molecular investigation of the tandem Tudor domain and plant homeodomain histone binding domains of the epigenetic regulator UHRF2

• Published in: Proteins, structure, function, and bioinformatics Vol. 90; no. 3; pp. 835 - 847
• Main Authors: Ginnard, Shane M., Winkler, Alyssa E., Mellado Fritz, Carlos, Bluhm, Tatum, Kemmer, Ray, Gilliam, Marisa, Butkevich, Nick, Abdrabbo, Sara, Bricker, Kaitlyn, Feiler, Justin, Miller, Isaak, Zoerman, Jenna, El‐Mohri, Zeineb, Khuansanguan, Panida, Basch, Madyson, Petzold, Timothy, Kostoff, Matthew, Konopka, Sean, Kociba, Brendon, Gillis, Thomas, Heyl, Deborah L., Trievel, Raymond C., Albaugh, Brittany N.
• Format: Journal Article
• Language: English
• Published: Hoboken, USA John Wiley & Sons, Inc 01.03.2022; Wiley Subscription Services, Inc
• Subjects: Affinity; Amino acids; Asparagine; Aspartic acid; Binding; binding interactions; Binding Sites; CCAAT-Enhancer-Binding Proteins - chemistry; chromatin; Crosstalk; Deoxyribonucleic acid; DNA; DNA Methylation; Electrostatic properties; Epigenesis, Genetic; Epigenetics; Escherichia coli - genetics; Gene Expression; Histone H3; Histones; Histones - chemistry; Homeobox; Homeodomain Proteins - chemistry; Humans; Ionic interactions; Melt temperature; Protein Binding; Protein Processing, Post-Translational; Proteins; Selectivity; structure comparison; Structure-Activity Relationship; Tudor Domain; Ubiquitin; Ubiquitin-Protein Ligases - chemistry; Ubiquitin-Protein Ligases - genetics; histones; binding interactions; chromatin; structure comparison; epigenetics
• ISSN: 0887-3585; 1097-0134; 1097-0134
• DOI: 10.1002/prot.26278

Ubiquitin‐like containing PHD and ring finger (UHRF)1 and UHRF2 are multidomain epigenetic proteins that play a critical role in bridging crosstalk between histone modifications and DNA methylation. Both proteins contain two histone reader domains, called tandem Tudor domain (TTD) and plant homeodomain (PHD), which read the modification status on histone H3 to regulate DNA methylation and gene expression. To shed light on the mechanism of histone binding by UHRF2, we have undergone a detailed molecular investigation with the TTD, PHD and TTD‐PHD domains and compared the binding activity to its UHRF1 counterpart. We found that unlike UHRF1 where the PHD is the primary binding contributor, the TTD of UHRF2 has modestly higher affinity toward the H3 tail, while the PHD has a weaker binding interaction. We also demonstrated that like UHRF1, the aromatic amino acids within the TTD are important for binding to H3K9me3 and a conserved aspartic acid within the PHD forms an ionic interaction with R2 of H3. However, while the aromatic amino acids in the TTD of UHRF1 contribute to selectivity, the analogous residues in UHRF2 contribute to both selectivity and affinity. We also discovered that the PHD of UHRF2 contains a distinct asparagine in the H3R2 binding pocket that lowers the binding affinity of the PHD by reducing a potential electrostatic interaction with the H3 tail. Furthermore, we demonstrate the PHD and TTD of UHRF2 cooperate to interact with the H3 tail and that dual domain engagement with the H3 tail relies on specific amino acids. Lastly, our data indicate that the unique stretch region in the TTD of UHRF2 can decrease the melting temperature of the TTD‐PHD and represents a disordered region. Thus, these subtle but important mechanistic differences are potential avenues for selectively targeting the histone binding interactions of UHRF1 and UHRF2 with small molecules.