DNA binding proteins that alter nucleic acid flexibility

Micah McCauley; Philip R. Hardwidge; L. James Maher III; Mark C. Williams

doi:10.1117/12.736306

5 September 2007 DNA binding proteins that alter nucleic acid flexibility

Micah McCauley, Philip R. Hardwidge, L. James Maher III, Mark C. Williams

Proceedings Volume 6644, Optical Trapping and Optical Micromanipulation IV; 664405 (2007) https://doi.org/10.1117/12.736306
Event: NanoScience + Engineering, 2007, San Diego, California, United States

Abstract

Dual - beam optical tweezers experiments subject single molecules of DNA to high forces (~ 300 pN) with 0.1 pN accuracy, probing the energy and specificity of nucleic acid - ligand structures. Stretching phage λ-DNA reveals an increase in the applied force up to a critical force known as the overstretching transition. In this region, base pairing and stacking are disrupted as double stranded DNA (dsDNA) is melted. Proteins that bind to the double strand will tend to stabilize dsDNA, and melting will occur at higher forces. Proteins that bind to single stranded DNA (ssDNA) destabilize melting, provided that the rate of association is comparable to the pulling rate of the experiment. Many proteins, however, exhibit some affinity for both dsDNA and ssDNA. We describe experiments upon DNA + HMGB2 (box A), a nuclear protein that is believed to facilitate transcription. By characterizing changes in the structure of dsDNA with a polymer model of elasticity, we have determined the equilibrium association constant for HMGB2 to be K_ds = 0.15 ± 0.7 10⁹ M^-1 for dsDNA binding. Analysis of the melting transition reveals an equilibrium association constant for HMGB2 to ssDNA to be K_ss = 0.039 ± 0.019 10⁹ M^-1 for ssDNA binding.

Citation Download Citation

Micah McCauley, Philip R. Hardwidge, L. James Maher III, and Mark C. Williams "DNA binding proteins that alter nucleic acid flexibility", Proc. SPIE 6644, Optical Trapping and Optical Micromanipulation IV, 664405 (5 September 2007); https://doi.org/10.1117/12.736306

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