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the melting of DNA

the melting of DNA. K = c A 2 / c A 2. For a self-complementary DNA sequence. 2 A ⇌ A 2. c 0 = c A + 2 c A 2. c A = c 0 – 2 c A 2. Total DNA strand concentration. K = c A 2 / (c 0 – 2 c A 2 ) 2. 4 Kc A 2 2 – (4 Kc 0 + 1) c A 2 + c 0 2 = 0. CHEM 471: Physical Chemistry.

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the melting of DNA

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  1. the melting of DNA K = cA2/cA2 For a self-complementary DNA sequence 2 A ⇌ A2 c0 = cA + 2cA2 cA = c0 – 2cA2 Total DNA strand concentration K = cA2/(c0 – 2cA2)2 4KcA2 2 – (4Kc0 + 1)cA2 + c02 = 0 CHEM 471: Physical Chemistry

  2. the melting of DNA At 50% melted K = 1/c0 Equilibrium constant for the melting reaction Km= 1/K = c0 ΔmG° = –RTmlnc0 = ΔmH° – TmΔmS°/R –ln c0 = ΔmH°/RTm – ΔmS°/R Tm = ΔmH°/(ΔmS° – R ln c0) DNA melting curves of (red) ATATATAT and (blue) GCGC CHEM 471: Physical Chemistry

  3. the melting of DNA DNA melting curves of (red) ATATATAT and (blue) GCGC Tm = ΔmH°/(ΔmS° – R ln c0) ΔfH°= –ΔmH° ΔfS° = –ΔmS° Tm = ΔfH°/(ΔfS° + R ln c0) CHEM 471: Physical Chemistry

  4. nearest neighbors and DNA thermodynamics Breslauer and co-workers found that if the next-nearest neighbors of the base pair were also considered, excellent and accurate values could be predicted for the enthalpy and entropy of melting of Watson-Crick hydrogen bonded ds-DNA of arbitrary sequence. Because GC base pairs have three hydrogen bonds to the two hydrogen bonds in AT pair, they are more stable, and so GC rich oligomers tend to melt at higher temperatures. • Modification by Santalucia (1997) • – Start with initiation step • – Add contributions from each pair of base pairs from table • — all values are for 1 M salt, 37 °C CHEM 471: Physical Chemistry

  5. nearest neighbors and DNA thermodynamics CHEM 471: Physical Chemistry

  6. nearest neighbors and DNA thermodynamics DNA sequence: ATAGGC Tm = ΔfH°/(ΔfS° + R ln c0) CHEM 471: Physical Chemistry ΔfH° = 0.8 + –(30.2 +30.2 +32.7+33.5+41.0) ΔfS° = –23.4 + –(85.4+89.2+87.9+83.3+102.2) Tm = (–166800 J/mol)/[–471.4 J mol–1K–1 + 8.3145 J mol–1K–1 × ln (.001)] = 354.9 K

  7. worth noting (1) CG/GC or GC/CG pairs of pairs, GG/CC not so much (2) G/C especially favorable in terminal position (3) Concentration matters more for short oligomers. (4) Melting curve requires calculating ln K = –ΔfH°/RTm + ΔfS°/R K = cA2/(c0 – 2cA2)2 (5) Note first order approximation to Taylor expansion of ΔfG° – ΔfC°p is unimportant! CHEM 471: Physical Chemistry (6) Symmetry correction to ΔfS°of +R ln 2 for self-complimentary duplexes

  8. ionic strength corrections ΔfS° = ΔfS° (1 M NaCl) + 1.540 J mol–1K–1 × N × ln [cNaCl] N is the number of base pairs in the duplex High salt balances the concentration of negative charges in the duplex, makes it more favorable. No significant enthalpic contribution CHEM 471: Physical Chemistry

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