Kinetics of the Triplex-Duplex Transition in DNA

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Kinetics of the Triplex-Duplex Transition in DNA Il-Buem Lee, Seok-Cheol Hong, Nam-Kyung Lee, Albert Johner  Biophysical Journal  Volume 103, Issue 12, Pages 2492-2501 (December 2012) DOI: 10.1016/j.bpj.2012.10.029 Copyright © 2012 Biophysical Society Terms and Conditions

Figure 1 (A) Triplex-forming DNA construct composed of a long mirror-repeat homo-pyrimidine sequence and a homo-purine sequence complementary to the 5′ side of the former. When the triplex folds, the energy transfer will occur between a donor (green, left-hatched) and an acceptor dye (red, solid). (B) Typical triplex-forming situation from a duplex DNA. Watson-Crick pairs (dashes). Hoogsteen pairs (stars). 3′ ends of the strands (gray arrowheads). In panels A and B, the triplex-forming (third) strand is shown (red base letters and red line, respectively). Biophysical Journal 2012 103, 2492-2501DOI: (10.1016/j.bpj.2012.10.029) Copyright © 2012 Biophysical Society Terms and Conditions

Figure 2 FRET efficiency histograms at various pH conditions. (A) 50 mM MES at pH 6.5, (B) 50 mM HEPES at pH 7.5, and (C) 50 mM Tris-HCl at pH 8.5. A triplex stabilized at low pH exhibits high FRET values. Biophysical Journal 2012 103, 2492-2501DOI: (10.1016/j.bpj.2012.10.029) Copyright © 2012 Biophysical Society Terms and Conditions

Figure 3 Calibration of the FRET efficiency with respect to the location of the donor dye. (A) The donor is next to the acceptor, (B) 6 nts away (HDNA-6), and (C) 9 nts away (HDNA-9). (Inset) Molecular arrangement for each measurement. The high FRET peak shifted according to the shift of donor position. Biophysical Journal 2012 103, 2492-2501DOI: (10.1016/j.bpj.2012.10.029) Copyright © 2012 Biophysical Society Terms and Conditions

Figure 4 The conformational distribution of the triplex-forming molecule and the (un)folding kinetics under various salt concentrations. Here pH was fixed to be 7.5 by 50 mM HEPES. The data in the first row (A, red, right-hatched) were obtained with [Na+] = 26 mM. The data in the second (B, blue, left-hatched) and the third (C, green, cross-hatched) rows show similar data with [Na+] = 50 and 100 mM, respectively. The first, second, third, and fourth columns show the FRET efficiency histograms, the dwell-time histograms of the low- and the high-FRET states, and representative FRET time traces of each corresponding condition, respectively. The time constants obtained for this set of data and other experimental data sets are summarized in Table S1. In the dwell-time distributions, we used a bin with the same size (0.1 s). Biophysical Journal 2012 103, 2492-2501DOI: (10.1016/j.bpj.2012.10.029) Copyright © 2012 Biophysical Society Terms and Conditions

Figure 5 (A) Optimal shape of the single strand of length S at the moment of contact. (B) Zipped configuration where the zipped section is indicated (dot-dashed line). (C) Optimal shape of the single-stranded loop at equilibrium with loop length S∗. The character O represents the origin of the XY coordinate system. In panels (B and C), the bound ssDNA strand is represented as if absorbed on the centerline of the duplex. Biophysical Journal 2012 103, 2492-2501DOI: (10.1016/j.bpj.2012.10.029) Copyright © 2012 Biophysical Society Terms and Conditions

Figure 6 A possible mechanism for spreading. (A) Before spreading. A loop inserted is located at the first (B) and second (C) base of the triplex. (Ladder) Duplex DNA. (Red circles) Nucleotides of the third strand. Biophysical Journal 2012 103, 2492-2501DOI: (10.1016/j.bpj.2012.10.029) Copyright © 2012 Biophysical Society Terms and Conditions