Hydrogen-deuterium exchange reveals a dynamic DNA-binding map of replication protein A
| Autor: | Brian Bothner, Faiz Ahmad, Edwin Antony, Angela Patterson, Jaigeeth Deveryshetty, Jenna Mattice, Nilisha Pokhrel |
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| Rok vydání: | 2020 |
| Předmět: |
Models
Molecular Protein Conformation AcademicSubjects/SCI00010 Allosteric regulation DNA Single-Stranded Hydrogen Deuterium Exchange-Mass Spectrometry Biology Genome Integrity Repair and Replication chemistry.chemical_compound Heterotrimeric G protein Replication Protein A Genetics Humans Replication protein A chemistry.chemical_classification Oligonucleotide DNA replication DNA-binding domain Oligosaccharide enzymes and coenzymes (carbohydrates) Enzyme chemistry Biophysics Hydrogen–deuterium exchange Linker DNA Protein Binding |
| Zdroj: | Nucleic Acids Research |
| ISSN: | 1362-4962 |
| Popis: | Replication protein A (RPA) binds to single-stranded DNA (ssDNA) and interacts with over three dozen enzymes and serves as a recruitment hub to coordinate most DNA metabolic processes. RPA binds ssDNA utilizing multiple oligosaccharide/oligonucleotide binding domains and based on their individual DNA binding affinities are classified as high versus low-affinity DNA-binding domains (DBDs). However, recent evidence suggests that the DNA-binding dynamics of DBDs better define their roles. Utilizing hydrogen–deuterium exchange mass spectrometry (HDX-MS), we assessed the ssDNA-driven dynamics of the individual domains of human RPA. As expected, ssDNA binding shows HDX changes in DBDs A, B, C, D and E. However, DBD-A and DBD-B are dynamic and do not show robust DNA-dependent protection. DBD-C displays the most extensive changes in HDX, suggesting a major role in stabilizing RPA on ssDNA. Slower allosteric changes transpire in the protein–protein interaction domains and linker regions, and thus do not directly interact with ssDNA. Within a dynamics-based model for RPA, we propose that DBD-A and -B act as the dynamic half and DBD-C, -D and -E function as the less-dynamic half. Thus, segments of ssDNA buried under the dynamic half are likely more readily accessible to RPA-interacting proteins. Graphical Abstract Graphical abstractRPA binds to DNA as two dynamic halves. |
| Databáze: | OpenAIRE |
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