TY - GEN
T1 - DNA binding proteins that alter nucleic acid flexibility
AU - McCauley, Micah
AU - Hardwidge, Philip R.
AU - Maher, L. James
AU - Williams, Mark C.
PY - 2007
Y1 - 2007
N2 - 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 X-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.
AB - 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 X-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.
KW - DNA protein binding
KW - Force induced melting
KW - Optical tweezers
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U2 - 10.1117/12.736306
DO - 10.1117/12.736306
M3 - Conference contribution
AN - SCOPUS:42149100440
SN - 9780819467928
T3 - Proceedings of SPIE - The International Society for Optical Engineering
BT - Optical Trapping and Optical Micromanipulation IV
T2 - Optical Trapping and Optical Micromanipulation IV
Y2 - 26 August 2007 through 29 August 2007
ER -