Kinetics and Thermodynamics of DNA, RNA, and Hybrid Duplex Formation

Rauzan, Brittany M.; McMichael, Elizabeth L.; Cave, Rachel; Sevcik, Lesley R; Ostrosky, Kara; Whitman, Elisabeth; Stegemann, Rachel; Sinclair, Audra L; Serra, Martin J.; Deckert, A. A.

doi:10.1021/bi3013005

Cited by 83 publications

(96 citation statements)

References 57 publications

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“…As the temperature was constant in our analysis, we infer that lnk s is proportional to the activation free energy, ΔG ‡ . The ΔG ‡ of the unfolding of a DNA duplex (ΔG ‡ off ) has a linear correlation with the stability of the DNA duplex (−ΔG°) (26), and the activation free energy of dissociation of a DNA duplex depends on the sequence (GC content) and length (27,28). Therefore, to unwind structured DNAs, the polymerase must decrease the ΔG ‡ off or rely on spontaneous dissociation of the structured DNA.…”

Section: Dependence Of Replication Efficiency On I-motif Stabilitymentioning

confidence: 99%

Topological impact of noncanonical DNA structures on Klenow fragment of DNA polymerase

Takahashi

Brazier

Sugimoto

2017

Proc. Natl. Acad. Sci. U.S.A.

113

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Noncanonical DNA structures that stall DNA replication can cause errors in genomic DNA. Here, we investigated how the noncanonical structures formed by sequences in genes associated with a number of diseases impacted DNA polymerization by the Klenow fragment of DNA polymerase. Replication of a DNA sequence forming an i-motif from a telomere, hypoxia-induced transcription factor, and an insulin-linked polymorphic region was effectively inhibited. On the other hand, replication of a mixed-type G-quadruplex (G4) from a telomere was less inhibited than that of the antiparallel type or parallel type. Interestingly, the i-motif was a better inhibitor of replication than were mixed-type G4s or hairpin structures, even though all had similar thermodynamic stabilities. These results indicate that both the stability and topology of structures formed in DNA templates impact the processivity of a DNA polymerase. This suggests that i-motif formation may trigger genomic instability by stalling the replication of DNA, causing intractable diseases.oncanonical intramolecular structures of nucleic acids, such as a triplex and a quadruplex, are stabilized under conditions that mimic the crowded cellular conditions (1), and have been detected in cells (2, 3). In vitro and in vivo, guaninequadruplex (G-quadruplex or G4) formation inhibits transcription and translation of template nucleic acids (4-8). It is possible that the noncanonical structures act as "functional codes" triggered by different molecular environments, which regulate gene expression epigenetically (6, 7). As sequences capable of forming the noncanonical structures are found in telomeres and promoter regions of known oncogenes, alterations of the noncanonical structures could play important roles in the progression of cancer and in other diseases (9).One of the remarkable features of noncanonical structures of nucleic acids is the diversity of topologies. In the case of G4s, antiparallel, mixed, and parallel type structures have been characterized ( Fig. 1 A-C). The sequences complementary to regions capable of G4 formation are composed of tandem repeats of cytosine, and these C-rich regions can form a different type of tetraplex topology, which is the i-motif (10, 11). An intramolecular i-motif is formed upon the interaction of four C-rich regions. The structure has three loops and two parallel-stranded hairpin-like units stabilized by cytosine and protonated cytosine (C:C + ) base pairs that are vertically intercalated antiparallel to one another ( Fig. 1 D and E). The i-motif structures are categorized into class I and class II based on the lengths of the loops. C-rich sequences are found in telomeres and in the promoter regions of about 40% of human genes (12, 13). As the i-motif is stabilized by hydrogen bonding in C:C + base pairs (14, 15), acidic conditions stabilize the structure. As the i-motif can mediate transcriptional regulation of B-cell lymphoma 2 (Bcl2) oncogene in cells (16), the environment inside these cells might be favorable to i-motif formation....

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Section: Dependence Of Replication Efficiency On I-motif Stabilitymentioning

confidence: 99%

Topological impact of noncanonical DNA structures on Klenow fragment of DNA polymerase

Takahashi

Brazier

Sugimoto

2017

Proc. Natl. Acad. Sci. U.S.A.

113

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“…Moreover, individual nucleotides within the RNA dendrimer can be labeled during chemical synthesis prior to dendrimer self-assembly, thus ensuring full utilization of void-spaces, which is an arduous challenge for small molecule based dendrimers. Compared to DNA counterparts, in addition to enhanced chemical stability after 2′-F modifications, RNA dendrimers are thermodynamically more stable 15,16,22,37,55,56 and will therefore not dissociate at ultra-low concentrations in vivo , and therapeutic RNA interference modules, such as siRNA and miRNA can be seamlessly integrated into the sequence design.…”

Section: Discussionmentioning

confidence: 99%

Controllable self-assembly of RNA dendrimers

Sharma

Haque

et al. 2016

Nanomedicine: Nanotechnology, Biology and Medicine

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We report programmable self-assembly of branched, 3D globular, monodisperse and nanoscale sized dendrimers using RNA as building blocks. The central core and repeating units of the RNA dendrimer are derivatives of the ultrastable three-way junction (3WJ) motif from the bacteriophage phi29 motor pRNA. RNA dendrimers were constructed by step-wise self assembly of modular 3WJ building blocks initiating with a single 3WJ core (Generation-0) with overhanging sticky-end and proceeding in a radial manner in layers up to Generation-4. The final constructs were generated under control without any structural defects in high yield and purity, as demonstrated by gel electrophoresis and AFM imaging. Upon incorporation of folate on the peripheral branches of the RNA dendrimers, the resulting constructs showed high binding and internalization into cancer cells. RNA dendrimers are envisioned to have a major impact in targeting, disease therapy, molecular diagnostics and bioelectronics in the near future.

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“…When there is no secondary structure to the single stranded DNA (e.g., hairpin structure), the rate constant, k on , will be ∼10 5 m -1 s -1 at 20-25°C with [Na + ] = 0.5-1 m (Hagan and Chakraborty 2004, Sekar et al 2005, Gao et al 2006, Rauzan et al 2013. Complementary strands hybridize virtually completely within 3 s at micromolar concentrations.…”

Section: Considerations For Probe Immobilization and Hybridization Stmentioning

confidence: 99%

Nucleic acid electrochemical and electromechanical biosensors: a review of techniques and developments

Rosario

Mutharasan

2014

Reviews in Analytical Chemistry

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This review comprises the last decade's development on experimental techniques for electrochemical and electromechanical sensing of nucleic acids, which originate from pathogenic bacteria, parasites, and viruses commonly found in food, water, and medical context. The electrochemical devices that are of primary interest are those that use voltammetry for detecting DNA and RNA-associated electrochemically active molecules at the working electrode. Attograms of nucleic acids have been reported to be detectable with electrochemical sensors in a batch-mode measurement arrangement. The masssensing electromechanical devices sense nucleic acids at the femtogram levels in a flow format without a molecular technique for amplifying target strand using polymerase chain reaction. Both underlying physics and methods of various studies are summarized, with discussion on limitations and potentials. We call attention to the need for sensors that not only detect but also confirm detection, as false negatives are not acceptable when one measures pathogenic species.

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Kinetics and Thermodynamics of DNA, RNA, and Hybrid Duplex Formation

Cited by 83 publications

References 57 publications

Topological impact of noncanonical DNA structures on Klenow fragment of DNA polymerase

Topological impact of noncanonical DNA structures on Klenow fragment of DNA polymerase

Controllable self-assembly of RNA dendrimers

Nucleic acid electrochemical and electromechanical biosensors: a review of techniques and developments

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