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Proceedings Paper

DNA binding proteins that alter nucleic acid flexibility
Author(s): Micah McCauley; Philip R. Hardwidge; L. James Maher; Mark C. Williams
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Paper 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 Kds = 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 Kss = 0.039 ± 0.019 109 M-1 for ssDNA binding.

Paper Details

Date Published: 5 September 2007
PDF: 12 pages
Proc. SPIE 6644, Optical Trapping and Optical Micromanipulation IV, 664405 (5 September 2007); doi: 10.1117/12.736306
Show Author Affiliations
Micah McCauley, Northeastern Univ. (United States)
Philip R. Hardwidge, Mayo Clinic College of Medicine (United States)
L. James Maher, Mayo Clinic College of Medicine (United States)
Mark C. Williams, Northeastern Univ. (United States)

Published in SPIE Proceedings Vol. 6644:
Optical Trapping and Optical Micromanipulation IV
Kishan Dholakia; Gabriel C. Spalding, Editor(s)

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