Fast Fenton Footprinting: A Laboratory-Based Method for the Time-Resolved Analysis of DNA, RNA and Proteins
'Footprinting' describes assays in which ligand binding or structure formation protects polymers such as nucleic acids and proteins from either cleavage or modification; footprinting allows the accessibility of individual residues to be mapped in solution. Equilibrium and time-dependent footprinting links site-specific structural information with thermodynamic and kinetic transitions. The hydroxyl radical ({center_dot}OH) is a particularly valuable footprinting probe by virtue of it being among the most reactive of chemical oxidants; it reports the solvent accessibility of reactive sites on macromolecules with as fine as a single residue resolution. A novel method of millisecond time-resolved {center_dot}OH footprinting has been developed based on the Fenton reaction, Fe(II) + H{sub 2}O{sub 2} {yields} Fe(III) + {center_dot}OH + OH{sup -}. This method can be implemented in laboratories using widely available three-syringe quench flow mixers and inexpensive reagents to study local changes in the solvent accessibility of DNA, RNA and proteins associated with their biological function.
- Research Organization:
- Brookhaven National Lab. (BNL), Upton, NY (United States). National Synchrotron Light Source
- Sponsoring Organization:
- Doe - Office Of Science
- DOE Contract Number:
- DE-AC02-98CH10886
- OSTI ID:
- 914346
- Report Number(s):
- BNL-78914-2007-JA; NARHAD; TRN: US200809%%188
- Journal Information:
- Nucleic Acids Res., Vol. 34; ISSN 0305-1048
- Country of Publication:
- United States
- Language:
- English
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