Investigations of the molecular crowding effect on nucleic acid structures and development of nucleotide switches

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The purposes of this study are construction of a database for nucleotide interactions and development of functional nucleotides that can be applied for a molecular sensing in the non-aqueous media such as in a cell and on a sensor surface. We found the DNA hybridization energy on a gold nanoparticle differed from that obtained in a homogeneous aqueous solution. A model system that mimics the molecular crowding environment supposedly occurring in a cell and on a gold nanoparticle was constructed by using PEG (polyethylene glycol) and other cosolutes. A large amount of cosolutes added in an aqueous solution substantially decreases the water activity and large-sized cosolutes generate large excluded volume effect. Investigations of oligonucleotide structures and their thermodynamic parameters in the cosolute-containing solutions revealed that the molecular crowding by PEG or other cosolute decreased the stability of Watson-Crick base pairs but stabilized the triplex, G-quadruplex, and lef … More t-handed duplex (Z-form) structures. To understand the molecular crowding effect, we analyzed the thermodynamic and kinetic parameters and obtained the notations that water molecules participating in a nucleotide structure formation significantly influence the thermodynamic stability and the kinetics under the molecular crowding conditions. According to the nucleotide interaction data, we developed DNA molecules that changed the conformation when binding to polyamines or basic peptides, and those formed a DNA-wire by transferring into the molecular crowding media. Moreover, we constructed the DNA molecules with a logic gate function that responded to the molecular environment. We also demonstrated that the molecular crowding effect was used for up-or down-regulation of an enzyme activity and an efficient PCR amplification of a GC-rich DNA sequence. We also successfully synthesized several types of functional nucleotides. The researches that focus on the molecular crowding are very important from the standpoint of medical and engineering aspects. Less