Salt has a biphasic effect on the higher-order structure of a DNA-protamine complex.

We observed single DNA molecules by fluorescence microscopy to clarify the effect of protamine on their higher-order structure. With an increase in the protamine concentration, the conformation of DNA molecules changes from an elongated coil state to a compact state through an intermediate state. Furthermore, the long-axis length of DNA gradually decreases while maintaining a distribution profile with a single peak. Such behavior is markedly different from the conformational transition of DNA induced by small polyamines such as spermidine and spermine, where individual DNA molecules exhibit an all-or-none transition from a coil to a globule state and the size distribution is characterized by twin peaks around the transition region. Next, we examined the effect of salt on the conformation of the DNA-protamine complex. Interestingly, at a fixed concentration of protamine, DNA tends to shrink with an increase in the NaCl concentration up to 300 mM, and then swells with a further increase in the NaCl concentration, that is, biphasic behavior is generated depending on the salt concentration. For comparison, we examined the effect of salt on DNA compaction induced by the trivalent polyamine spermidine. We confirmed that salt always has an inhibitory effect on spermine-induced compaction. To clarify this biphasic effect of salt on protamine-induced DNA compaction, we performed a numerical simulation on a negatively charged semiflexible polyelectrolyte in the presence of polycations with relatively large numbers of positive charges by taking into account the effect of salt at different concentrations. The results showed that salt promotes compaction up to a certain concentration and then tends to unfold the polyelectrolyte chain, which reproduced the experimental observation in a semiquantitative manner. This biphasic effect is discussed in relation to the specific shielding effect that depends on the salt concentration.

Thermal modulation voltammetry at a 1,2-dichloroethane/water microinterface using visible laser heating with optically absorbing supporting electrolyte.

Thermal modulation voltammetry (TMV) using laser heating at a liquid/liquid microinterface is a useful technique for determining the standard entropy change of ion transfer. In this work, for achieving TMV using a visible laser as a heating source at a 1,2-dichloroethane (DCE)/water microinterface, we used an optically absorbing supporting electrolyte, i.e., crystalviolet tetrakis(4-chlorophenyl)borate (CVTClPB). CVTClPB served well not only as a supporting electrolyte but also as an optical absorber in the DCE solution, and thereby, well-defined linear sweep (LS) and TM voltammograms could be obtained. On the basis of LS and thermal modulation (TM) voltammograms obtained with CVTClPB, the standard entropy changes of ion transfer for six model ions were successfully determined.

Folding transition of a single semiflexible polyelectrolyte chain through toroidal bundling of loop structures.

We consider how the DNA coil-globule transition progresses via the formation of a toroidal ring structure. We formulate a theoretical model of this transition as a phenomenon in which an unstable single loop generated as a result of thermal fluctuation is stabilized through association with other loops along a polyelectrolyte chain. An essential property of the chain under consideration is that it follows a wormlike chain model. A toroidal bundle of loop structures is characterized by a radius and a winding number. The statistical properties of such a chain are discussed in terms of the free energy as a function of the fraction of unfolded segments. We also present an actual experimental observation of the coil-globule transition of single giant DNA molecules, T4 DNA (165.5 kbp), with spermidine (3+), where intrachain phase segregation appears at a NaCl concentration of more than 10 mM. Both the theory and experiments lead to two important points. First, the transition from a partially folded state to a completely folded state has the characteristics of a continuous transition, while the transition from an unfolded state to a folded state has the characteristics of a first-order phase transition. Second, the appearance of a partially folded structure requires a folded structure to be less densely packed than in the fully folded compact state.

NTP concentration switches transcriptional activity by changing the large-scale structure of DNA.

It is becoming clearer that genetic activity is closely associated with the intracellular energy state. However, the mechanisms of this association are still unclear. In this study, we focused on large-scale changes in the structure of DNA to examine the effect of the NTP concentration on the transcription reaction with T7 RNA polymerase and compared the results with long duplex DNA to those with a short persistent-length(1) fragment. The transcriptional activity dramatically changed only for long duplex DNA within a narrow range of NTP concentrations associated with changes in the large-scale structure of DNA. This result suggests that the energy state may play an essential role in regulating ON/OFF switching on transcriptional activity.

Proton concentration (pH) switches the higher-order structure of DNA in the presence of spermine.

Single-chain observations on the conformational change of giant DNA (T4 DNA) molecules were performed using fluorescence microscopy at different values of pH in the presence of spermine. Individual DNA molecules undergo a large discrete change, or all-or-none transition, in conformation from a folded compact state to an unfolded coil state with an increase in pH. This abrupt unfolding of DNA with an increase in pH is attributed to a decrease in the concentration of the tetravalent form in spermine [SPM(4+)]. We propose a scheme for the folding transition of single DNAs, where the manner of spermine binding changes dramatically from weak loose binding in the elongated coil state to strong tight binding in the folded compact state. We discuss the hierarchical nature of the transition, i.e. cooperative continuous change on the ensemble vs. all-or-none switching on individual DNAs.