TY - JOUR
T1 - Conformational energetics of stable and metastable states formed by DNA triplet repeat oligonucleotides
T2 - Implications for triplet expansion diseases
AU - Völker, J.
AU - Makube, N.
AU - Plum, G. E.
AU - Klump, H. H.
AU - Breslauer, K. J.
PY - 2002/11/12
Y1 - 2002/11/12
N2 - We have embedded the hexameric triplet repeats (CAG)6 and (CTG)6 between two (GC)3 domains to produce two 30-mer hairpins with the sequences d[(GC)3(CAG)6(GC)3] and d[(GC)3(CTG)6(GC)3]. This construct reduces the conformational space available to these repetitive DNA sequences. We find that the (CAG)6 and (CTG)6 repeats form stable, ordered, single-stranded structures. These structures are stabilized at 62°C by an average enthalpy per base of 1.38 kcal·mol-1 for the CAG triplet and 2.87 kcal·mol-1 for the CTG triplet, while being entropically destabilized by 3.50 cal·K-1·mol-1 for the CAG triplet and 7.6 cal·K-1·mol-1 for the CTG triplet. Remarkably, these values correspond, respectively, to 1/3 (for CAG) and 2/3 (for CTG) of the enthalpy and entropy per base values associated with Watson-Crick base pairs. We show that the presence of the loop structure kinetically inhibits duplex formation from the two complementary 30-mer hairpins, even though the duplex is the thermodynamically more stable state. Duplex formation, however, does occur at elevated temperatures. We propose that this thermally induced formation of a more stable duplex results from thermal disruption of the single-stranded order, thereby allowing the complementary domains to associate (perhaps via "kissing hairpins"). Our melting profiles show that, once duplex formation has occurred, the hairpin intermediate state cannot be reformed, consistent with our interpretation of kinetically trapped hairpin structures. The duplex formed by the two complementary oligonucleotides does not have any unusual optical or thermodynamic properties. By contrast, the very stable structures formed by the individual single-stranded triplet repeat sequences are thermally and thermodynamically unusual. We discuss this stable, triplet repeat, single-stranded structure and its interconversion with duplex in terms of triplet expansion diseases.
AB - We have embedded the hexameric triplet repeats (CAG)6 and (CTG)6 between two (GC)3 domains to produce two 30-mer hairpins with the sequences d[(GC)3(CAG)6(GC)3] and d[(GC)3(CTG)6(GC)3]. This construct reduces the conformational space available to these repetitive DNA sequences. We find that the (CAG)6 and (CTG)6 repeats form stable, ordered, single-stranded structures. These structures are stabilized at 62°C by an average enthalpy per base of 1.38 kcal·mol-1 for the CAG triplet and 2.87 kcal·mol-1 for the CTG triplet, while being entropically destabilized by 3.50 cal·K-1·mol-1 for the CAG triplet and 7.6 cal·K-1·mol-1 for the CTG triplet. Remarkably, these values correspond, respectively, to 1/3 (for CAG) and 2/3 (for CTG) of the enthalpy and entropy per base values associated with Watson-Crick base pairs. We show that the presence of the loop structure kinetically inhibits duplex formation from the two complementary 30-mer hairpins, even though the duplex is the thermodynamically more stable state. Duplex formation, however, does occur at elevated temperatures. We propose that this thermally induced formation of a more stable duplex results from thermal disruption of the single-stranded order, thereby allowing the complementary domains to associate (perhaps via "kissing hairpins"). Our melting profiles show that, once duplex formation has occurred, the hairpin intermediate state cannot be reformed, consistent with our interpretation of kinetically trapped hairpin structures. The duplex formed by the two complementary oligonucleotides does not have any unusual optical or thermodynamic properties. By contrast, the very stable structures formed by the individual single-stranded triplet repeat sequences are thermally and thermodynamically unusual. We discuss this stable, triplet repeat, single-stranded structure and its interconversion with duplex in terms of triplet expansion diseases.
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U2 - 10.1073/pnas.222519799
DO - 10.1073/pnas.222519799
M3 - Article
C2 - 12417759
AN - SCOPUS:0037069430
SN - 0027-8424
VL - 99
SP - 14700
EP - 14705
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 23
ER -