Extra Adenosine Stacks into the Self-Complementary d(CGCAGAATTCGCG) Duplex in Solution

Dinshaw J. Patel, Sharon A. Kozlowski, Janet A. Rice, Chris Broka, Keiichi Itakura, Luis A. Marky, Kenneth Breslauer

Research output: Contribution to journalArticle

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Abstract

The structural and energetic parameters for the helix to coil transition of the duplex formed by the self-complementary d(C1G2C3AG4A5A6T6T5C4G3C2G1) sequence (henceforth called 13-mer duplex) that contains an extra noncomplementary dA residue between positions dC3 and dG4 are compared with the duplex formed by the corresponding self-complementary d(CGCGAATTCGCG) sequence (henceforth called 12-mer duplex) that lacks the extra base [Patel, D.J., Kozlowski, S.A., Marky, L.A., Broka, C., Rice, J.A., Itakuru, K., & Breslauer, K.J. (1981) Biochemistry (first paper of four in this issue)]. The nonexchangeable base proton nuclear magnetic resonance (NMR) resonances from the extra dA residue are identified and monitored through the melting transition of the 13-mer duplex. These markers demonstrate that the adenosine stacks into the 13-mer duplex rather than form a bulge loop. Furthermore, the base pairing on either side of the extra stacked dA residue remains intact. A downfield-shifted phosphorus resonance in the 31P spectrum of the 13-mer duplex is tentatively assigned to the extended phosphodiester linkage opposite the stacked, extra dA residue. The nonexchangeable protons from base pairs 2–6 and the extra dA residue exhibit a duplex to strand transition midpoint of 57 ± 2 °C in 0.1 M phosphate solution. The thermally induced transition of the 13-mer duplex was also monitored by differential scanning calorimetry. This technique reveals that insertion of an extra dA into each strand of the duplex results in a 19 °C reduction in the melting temperature of the 13-mer relative to the 12-mer duplex. A calorimetric enthalpy change of 104 kcal (mol of double strand)−1 and a van't Hoff enthalpy change of 70 kcal were determined for the helix to coil transition of the 13-mer duplex in 0.1 M NaCl solution. Comparison of these enthalpies reveals that the transition is not a two-state process and that the size of the cooperative unit is 9 ± 1 base pairs. The similarity between the calorimetric transition enthalpies for the 12-mer and 13-mer duplexes is consistent with a picture in which the stacking of the extra dA residue into the duplex on one strand compensates for the loss of stacking between dG·dC base pairs 3 and 4 at the site of the helix interruption.

Original languageEnglish (US)
Pages (from-to)445-451
Number of pages7
JournalBiochemistry
Volume21
Issue number3
DOIs
StatePublished - Feb 1 1982

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Base Pairing
Adenosine
Enthalpy
Freezing
Protons
Nuclear magnetic resonance
Biochemistry
Differential Scanning Calorimetry
Phosphorus
Melting point
Differential scanning calorimetry
Melting
Magnetic Resonance Spectroscopy
Phosphates
Temperature

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Patel, D. J., Kozlowski, S. A., Rice, J. A., Broka, C., Itakura, K., Marky, L. A., & Breslauer, K. (1982). Extra Adenosine Stacks into the Self-Complementary d(CGCAGAATTCGCG) Duplex in Solution. Biochemistry, 21(3), 445-451. https://doi.org/10.1021/bi00532a004
Patel, Dinshaw J. ; Kozlowski, Sharon A. ; Rice, Janet A. ; Broka, Chris ; Itakura, Keiichi ; Marky, Luis A. ; Breslauer, Kenneth. / Extra Adenosine Stacks into the Self-Complementary d(CGCAGAATTCGCG) Duplex in Solution. In: Biochemistry. 1982 ; Vol. 21, No. 3. pp. 445-451.
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abstract = "The structural and energetic parameters for the helix to coil transition of the duplex formed by the self-complementary d(C1G2C3AG4A5A6T6T5C4G3C2G1) sequence (henceforth called 13-mer duplex) that contains an extra noncomplementary dA residue between positions dC3 and dG4 are compared with the duplex formed by the corresponding self-complementary d(CGCGAATTCGCG) sequence (henceforth called 12-mer duplex) that lacks the extra base [Patel, D.J., Kozlowski, S.A., Marky, L.A., Broka, C., Rice, J.A., Itakuru, K., & Breslauer, K.J. (1981) Biochemistry (first paper of four in this issue)]. The nonexchangeable base proton nuclear magnetic resonance (NMR) resonances from the extra dA residue are identified and monitored through the melting transition of the 13-mer duplex. These markers demonstrate that the adenosine stacks into the 13-mer duplex rather than form a bulge loop. Furthermore, the base pairing on either side of the extra stacked dA residue remains intact. A downfield-shifted phosphorus resonance in the 31P spectrum of the 13-mer duplex is tentatively assigned to the extended phosphodiester linkage opposite the stacked, extra dA residue. The nonexchangeable protons from base pairs 2–6 and the extra dA residue exhibit a duplex to strand transition midpoint of 57 ± 2 °C in 0.1 M phosphate solution. The thermally induced transition of the 13-mer duplex was also monitored by differential scanning calorimetry. This technique reveals that insertion of an extra dA into each strand of the duplex results in a 19 °C reduction in the melting temperature of the 13-mer relative to the 12-mer duplex. A calorimetric enthalpy change of 104 kcal (mol of double strand)−1 and a van't Hoff enthalpy change of 70 kcal were determined for the helix to coil transition of the 13-mer duplex in 0.1 M NaCl solution. Comparison of these enthalpies reveals that the transition is not a two-state process and that the size of the cooperative unit is 9 ± 1 base pairs. The similarity between the calorimetric transition enthalpies for the 12-mer and 13-mer duplexes is consistent with a picture in which the stacking of the extra dA residue into the duplex on one strand compensates for the loss of stacking between dG·dC base pairs 3 and 4 at the site of the helix interruption.",
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Patel, DJ, Kozlowski, SA, Rice, JA, Broka, C, Itakura, K, Marky, LA & Breslauer, K 1982, 'Extra Adenosine Stacks into the Self-Complementary d(CGCAGAATTCGCG) Duplex in Solution', Biochemistry, vol. 21, no. 3, pp. 445-451. https://doi.org/10.1021/bi00532a004

Extra Adenosine Stacks into the Self-Complementary d(CGCAGAATTCGCG) Duplex in Solution. / Patel, Dinshaw J.; Kozlowski, Sharon A.; Rice, Janet A.; Broka, Chris; Itakura, Keiichi; Marky, Luis A.; Breslauer, Kenneth.

In: Biochemistry, Vol. 21, No. 3, 01.02.1982, p. 445-451.

Research output: Contribution to journalArticle

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T1 - Extra Adenosine Stacks into the Self-Complementary d(CGCAGAATTCGCG) Duplex in Solution

AU - Patel, Dinshaw J.

AU - Kozlowski, Sharon A.

AU - Rice, Janet A.

AU - Broka, Chris

AU - Itakura, Keiichi

AU - Marky, Luis A.

AU - Breslauer, Kenneth

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N2 - The structural and energetic parameters for the helix to coil transition of the duplex formed by the self-complementary d(C1G2C3AG4A5A6T6T5C4G3C2G1) sequence (henceforth called 13-mer duplex) that contains an extra noncomplementary dA residue between positions dC3 and dG4 are compared with the duplex formed by the corresponding self-complementary d(CGCGAATTCGCG) sequence (henceforth called 12-mer duplex) that lacks the extra base [Patel, D.J., Kozlowski, S.A., Marky, L.A., Broka, C., Rice, J.A., Itakuru, K., & Breslauer, K.J. (1981) Biochemistry (first paper of four in this issue)]. The nonexchangeable base proton nuclear magnetic resonance (NMR) resonances from the extra dA residue are identified and monitored through the melting transition of the 13-mer duplex. These markers demonstrate that the adenosine stacks into the 13-mer duplex rather than form a bulge loop. Furthermore, the base pairing on either side of the extra stacked dA residue remains intact. A downfield-shifted phosphorus resonance in the 31P spectrum of the 13-mer duplex is tentatively assigned to the extended phosphodiester linkage opposite the stacked, extra dA residue. The nonexchangeable protons from base pairs 2–6 and the extra dA residue exhibit a duplex to strand transition midpoint of 57 ± 2 °C in 0.1 M phosphate solution. The thermally induced transition of the 13-mer duplex was also monitored by differential scanning calorimetry. This technique reveals that insertion of an extra dA into each strand of the duplex results in a 19 °C reduction in the melting temperature of the 13-mer relative to the 12-mer duplex. A calorimetric enthalpy change of 104 kcal (mol of double strand)−1 and a van't Hoff enthalpy change of 70 kcal were determined for the helix to coil transition of the 13-mer duplex in 0.1 M NaCl solution. Comparison of these enthalpies reveals that the transition is not a two-state process and that the size of the cooperative unit is 9 ± 1 base pairs. The similarity between the calorimetric transition enthalpies for the 12-mer and 13-mer duplexes is consistent with a picture in which the stacking of the extra dA residue into the duplex on one strand compensates for the loss of stacking between dG·dC base pairs 3 and 4 at the site of the helix interruption.

AB - The structural and energetic parameters for the helix to coil transition of the duplex formed by the self-complementary d(C1G2C3AG4A5A6T6T5C4G3C2G1) sequence (henceforth called 13-mer duplex) that contains an extra noncomplementary dA residue between positions dC3 and dG4 are compared with the duplex formed by the corresponding self-complementary d(CGCGAATTCGCG) sequence (henceforth called 12-mer duplex) that lacks the extra base [Patel, D.J., Kozlowski, S.A., Marky, L.A., Broka, C., Rice, J.A., Itakuru, K., & Breslauer, K.J. (1981) Biochemistry (first paper of four in this issue)]. The nonexchangeable base proton nuclear magnetic resonance (NMR) resonances from the extra dA residue are identified and monitored through the melting transition of the 13-mer duplex. These markers demonstrate that the adenosine stacks into the 13-mer duplex rather than form a bulge loop. Furthermore, the base pairing on either side of the extra stacked dA residue remains intact. A downfield-shifted phosphorus resonance in the 31P spectrum of the 13-mer duplex is tentatively assigned to the extended phosphodiester linkage opposite the stacked, extra dA residue. The nonexchangeable protons from base pairs 2–6 and the extra dA residue exhibit a duplex to strand transition midpoint of 57 ± 2 °C in 0.1 M phosphate solution. The thermally induced transition of the 13-mer duplex was also monitored by differential scanning calorimetry. This technique reveals that insertion of an extra dA into each strand of the duplex results in a 19 °C reduction in the melting temperature of the 13-mer relative to the 12-mer duplex. A calorimetric enthalpy change of 104 kcal (mol of double strand)−1 and a van't Hoff enthalpy change of 70 kcal were determined for the helix to coil transition of the 13-mer duplex in 0.1 M NaCl solution. Comparison of these enthalpies reveals that the transition is not a two-state process and that the size of the cooperative unit is 9 ± 1 base pairs. The similarity between the calorimetric transition enthalpies for the 12-mer and 13-mer duplexes is consistent with a picture in which the stacking of the extra dA residue into the duplex on one strand compensates for the loss of stacking between dG·dC base pairs 3 and 4 at the site of the helix interruption.

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Patel DJ, Kozlowski SA, Rice JA, Broka C, Itakura K, Marky LA et al. Extra Adenosine Stacks into the Self-Complementary d(CGCAGAATTCGCG) Duplex in Solution. Biochemistry. 1982 Feb 1;21(3):445-451. https://doi.org/10.1021/bi00532a004