Protein–DNA and ion–DNA interactions revealed through contrast variation SAXS
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Understanding how DNA carries out its biological roles requires knowledge of its interactions with biological partners. Since DNA is a polyanionic polymer, electrostatic interactions contribute significantly. These interactions are mediated by positively charged protein residues or charge compensating cations. Direct detection of these partners and/or their effect on DNA conformation poses challenges, especially for monitoring conformational dynamics in real time. Small-angle x-ray scattering (SAXS) is uniquely sensitive to both the conformation and local environment (i.e. protein partner and associated ions) of the DNA. The primary challenge of studying multi-component systems with SAXS lies in resolving how each component contributes to the measured scattering. Here, we review two contrast variation (CV) strategies that enable targeted studies of the structures of DNA or its associated partners. First, solution contrast variation enables measurement of DNA conformation within a protein–DNA complex by masking out the protein contribution to the scattering profile. We review a specific example, in which the real-time unwrapping of DNA from a nucleosome core particle is measured during salt-induced disassembly. The second method, heavy atom isomorphous replacement, reports the spatial distribution of the cation cloud around duplex DNA by exploiting changes in the scattering strength of cations with varying atomic numbers. We demonstrate the application of this approach to provide the spatial distribution of monovalent cations (Na+, K+, Rb+, Cs+) around a standard 25-base pair DNA. The CV strategies presented here are valuable tools for understanding DNA interactions with its biological partners.
KeywordsSAXS Contrast variation NCP DNA Ions Heavy atom isomorphous replacement
The work was supported by National Institutes of Health grants (EUREKA R01-GM088645) and (R01-GM085062). J.M.T. was supported by National Institutes of Health Training Grant (T32GM008267).
Compliance with ethical standards
Conflict of interest
Joshua M. Tokuda declares that he has no conflict of interest.
Suzette A. Pabit declares that she has no conflict of interest.
Lois Pollack declares that she has no conflict of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
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