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Daisuke Miyoshi

Publications and source records attributed to Daisuke Miyoshi.

23 records · Page 2Linked to original sources
PubMed2003

Structural transition from antiparallel to parallel G-quadruplex of d(G4T4G4) induced by Ca2+.

Guanine quadruplex (G-quadruplex) structures are formed by guanine-rich oligonucleotides. Because of their in vivo and in vitro importance, numerous studies have been demonstrated that the structure and stability of the G-quadruplex are dependent on the sequence of oligonucleotide and environmental conditions such as existing cations. Previously, we quantitatively investigated the divalent cation effects on the antiparallel G-quadruplex of d(G4T4G4), and found that Ca2+ induces a structural transition from the antiparallel to parallel G-quadruplex, and finally G-wire formation. In the present study, we report in detail the kinetic and thermodynamic analyses of the structural transition induced by Ca2+ using stopped-flow apparatus, circular dichroism, size-exclusion chromatography (SEC) and atomic force microscopy. The quantitative parameters showed that at least two Ca2+ ions were required for the transition. The kinetic parameters also indicated that d(G4T4G4) underwent the transition through multiple steps involving the Ca2+ binding, isomerization and oligomerization of d(G4T4G4). The parallel-stranded G-wire structure of d(G4T4G4), which is a well controlled alignment of numerous DNA strands with G-quartets, as the final product induced by Ca2+, was observed using SEC and atomic force microscopy. These results provide insight into the mechanism of the structural transition and G-wire formation and are useful for constructing a nanomaterial regulated by Ca2+.

Base Sequence↗
PubMed2003

Structural polymorphism of telomeric DNA regulated by pH and divalent cation.

DNA oligonucleotides can form multi-stranded structures such as a duplex, triplex, and quadruplex, while the double helical structure is generally considered as the canonical structure of DNA oligonucleotides. Guanine-rich or cytosine-rich oligonucleotides, which are observed in telomere, centromere, and other biologically important sequences in vivo, can form four-stranded G-quadruplex and I-motif structures in vitro. In this study, we have investigated the effects of pH and cation on the structures and their stabilities of d(G4T4G4) and d(C4A4C4). The CD spectra and thermal melting curves of DNAs at various pHs demonstrated that acidic conditions induced a stable I-motif structure of d(C4A4C4), while the pH value did not affect the G-quadruplex structure and stability of d(G4T4G4). The CD spectra of the 1:1 mixture of d(G4T4G4) and d(C4A4C4) indicated that the acidic conditions inhibit the duplex formation between d(G4T4G4) and d(C4A4C4). Isothermal titration calorimetry measurements of the duplex formation at various pHs also quantitatively indicated that the acidic conditions inhibit the duplex formation. On the other hand, the CD spectra and thermal melting curves of DNAs in the absence and presence of Ca2+ indicated that Ca2+ induces a parallel G-quadruplex structure of d(G4T4G4) and then inhibits the duplex formation. These results lead to the conclusion that both the pH and coexisting cation can induce and regulate the structural polymorphisms the oligonucleotides in which they form the G-quadruplex, I-motif, and duplex depending on the conditions. Thus, the results reported here indicate pivotal roles of pH and coexisting cations in biological processes by regulating the conformational switching between the duplex and quadruplexes structures of the guanine-rich or cytosine-rich oligonucleotides in vivo.

Cations, Divalent↗
PubMed2002

Molecular crowding regulates the structural switch of the DNA G-quadruplex.

Almost all biochemical reactions in vitro have been investigated through numerous experiments conducted in dilute solutions containing low concentrations of solutes. However, biomacromolecules such as nucleic acids, proteins, and polysaccharides are designed to function and/or form their native structures in a living cell containing high concentrations of biomacromolecules, substrates, cofactors, salts, and so on. In the present study, we have demonstrated quantitatively the effect of molecular crowding on structures and stabilities of the G-quadruplex of d(G(4)T(4)G(4)). Molecular crowding with poly(ethylene glycol) (PEG) induced a structural transition from the antiparallel to the parallel G-quadruplex of d(G(4)T(4)G(4)), while molecular crowding with polycations did not alter the structure of the antiparallel G-quadruplex. The binding constants of putrescine, one of the polycations, for d(G(4)T(4)G(4)) in the absence and presence of Na(+) are calculated to be 277 and 2.5 M(-)(1), respectively. This indicates that the polycations coordinate to d(G(4)T(4)G(4)) with electrostatic interactions. The thermodynamic parameters of the antiparallel G-quadruplex formation under the crowding and noncrowding conditions induced by putrescine were also estimated. The stability of the antiparallel G-quadruplex decreased (-DeltaG degrees (25) decreased from 28 to 22 kcal mol(-)(1)) with molecular crowding by putrescine. Also, enthalpy and entropy changes in the structural formation under crowding and noncrowding conditions clearly showed that destabilization was entropy-driven. These quantitative parameters indicated that both the volume excluded by PEG and chemical interactions such as electrostatic interaction with solute polycations are critical for determining how molecular crowding affects the structure and stability of highly ordered DNA structures.

Animals↗
PubMed2002

Long RNA dangling end has large energetic contribution to duplex stability.

Long terminal unpaired nucleotides known as dangling ends play interesting roles in biological systems. Previous studies, however, only dealt with the energy contributions of single dangling bases. The energy contributions of long dangling ends on the stability of duplexes have not been systematically studied. We now report a quantitative increase in stability of RNA-RNA and DNA-DNA duplexes containing a long dangling end. We found a larger enhancement of the stability by the long RNA dangling end of the RNA-RNA duplex than has been observed for the DNA duplexes. It is also found that structural stabilizations by long dangling ends seem to originate from the single-stranded stacking interactions of nucleotides. These results indicate that RNA stability can be achieved by increasing the length of the dangling end. The thermodynamic parameters of the long dangling ends are useful for designing ribozymes and antisense oligonucleotides, and for the prediction of the RNA secondary structure like the pseudoknot.

DNA↗