Use of Nucleic Acid Analogs for the Study of Nucleic Acid Interactions

Unnatural nucleosides have been explored to expand the properties and the applications of oligonucleotides. This paper briefly summarizes nucleic acid analogs in which the base is modified or replaced by an unnatural stacking group for the study of nucleic acid interactions. We also describe the nucleoside analogs of a base pair-mimic structure that we have examined. Although the base pair-mimic nucleosides possess a simplified stacking moiety of a phenyl or naphthyl group, they can be used as a structural analog of Watson-Crick base pairs. Remarkably, they can adopt two different conformations responding to their interaction energies, and one of them is the stacking conformation of the nonpolar aromatic group causing the site-selective flipping of the opposite base in a DNA double helix. The base pair-mimic nucleosides can be used to study the mechanism responsible for the base stacking and the flipping of bases out of a nucleic acid duplex

The Effects of Molecular Crowding on the Structure and Stability of G-Quadruplexes with an Abasic Site

Both cellular environmental factors and chemical modifications critically affect the properties of nucleic acids. However, the structure and stability of DNA containing abasic sites under cell-mimicking molecular crowding conditions remain unclear. Here, we investigated the molecular crowding effects on the structure and stability of the G-quadruplexes including a single abasic site. Structural analysis by circular dichroism showed that molecular crowding by PEG200 did not affect the topology of the G-quadruplex structure with or without an abasic site. Thermodynamic analysis further demonstrated that the degree of stabilization of the G-quadruplex by molecular crowding decreased with substitution of an abasic site for a single guanine. Notably, we found that the molecular crowding effects on the enthalpy change for G-quadruplex formation had a linear relationship with the abasic site effects depending on its position. These results are useful for predicting the structure and stability of G-quadruplexes with abasic sites in the cell-mimicking conditions

RNA/DNA hybrid duplexes with identical nearest-neighbor base-pairs have identical stability

AbstractEnergetic behaviors of eight pairs of RNA/DNA hybrid duplexes with identical nearest neighbors have been investigated by UV melting analysis. In the pairs with identical nearest-neighbor pairs, the melting curve traces at the same strand concentration were very similar. The average difference in stabilization energy of these pairs was 4%, which was about expected within experimental error. These results indicate that the nearest-neighbor model is valid for predicting the stability of RNA/DNA hybrid duplexes as well as RNA/RNA and DNA/DNA duplexes

DNA base flipping by a base pair-mimic nucleoside

On the basis of non-covalent bond interactions in nucleic acids, we synthesized the deoxyadenosine derivatives tethering a phenyl group (X) and a naphthyl group (Z) by an amide linker, which mimic a Watson–Crick base pair. Circular dichroism spectra indicated that the duplexes containing X and Z formed a similar conformation regardless of the opposite nucleotide species (A, G, C, T and an abasic site analogue F), which was not observed for the natural duplexes. The [Formula: see text] values among the natural duplexes containing the A/A, A/G, A/C, A/T and A/F pairs differed by 5.2 kcal mol(−1) while that among the duplexes containing X or Z in place of the adenine differed by only 1.9 or 2.8 kcal mol(−1), respectively. Fluorescence quenching experiments confirmed that 2-amino purine opposite X adopted an unstacked conformation. The structural and thermodynamic analyses suggest that the aromatic hydrocarbon group of X and Z intercalates into a double helix, resulting in the opposite nucleotide base flipping into an unstacked position regardless of the nucleotide species. This observation implies that modifications at the aromatic hydrocarbon group and the amide linker may expand the application of the base pair-mimic nucleosides for molecular biology and biotechnology

Hydration Changes upon DNA Folding Studied by Osmotic Stress Experiments

AbstractThe thermal stability of nucleic acid structures is perturbed under the conditions that mimic the intracellular environment, typically rich in inert components and under osmotic stress. We now describe the thermodynamic stability of DNA oligonucleotide structures in the presence of high background concentrations of neutral cosolutes. Small cosolutes destabilize the basepair structures, and the DNA structures consisting of the same nearest-neighbor composition show similar thermodynamic parameters in the presence of various types of cosolutes. The osmotic stress experiments reveal that water binding to flexible loops, unstable mismatches, and an abasic site upon DNA folding are almost negligible, whereas the binding to stable mismatch pairs is significant. The studies using the basepair-mimic nucleosides and the peptide nucleic acid suggest that the sugar-phosphate backbone and the integrity of the basepair conformation make important contributions to the binding of water molecules to the DNA bases and helical grooves. The study of the DNA hydration provides the basis for understanding and predicting nucleic acid structures in nonaqueous solvent systems

Newly characterized interaction stabilizes DNA structure: oligoethylene glycols stabilize G-quadruplexes CH–π interactions

Oligoethylene glycols are used as crowding agents in experiments that aim to understand the effects of intracellular environments on DNAs. Moreover, DNAs with covalently attached oligoethylene glycols are used as cargo carriers for drug delivery systems. To investigate how oligoethylene glycols interact with DNAs, we incorporated deoxythymidine modified with oligoethylene glycols of different lengths, such as tetraethylene glycol (TEG), into DNAs that form antiparallel G-quadruplex or hairpin structures such that the modified residues were incorporated into loop regions. Thermodynamic analysis showed that because of enthalpic differences, the modified G-quadruplexes were stable and the hairpin structures were slightly unstable relative to unmodified DNA. The stability of G-quadruplexes increased with increasing length of the ethylene oxides and the number of deoxythymidines modified with ethylene glycols in the G-quadruplex. Nuclear magnetic resonance analyses and molecular dynamics calculations suggest that TEG interacts with bases in the G-quartet and loop via CH-pi and lone pair-pi interactions, although it was previously assumed that oligoethylene glycols do not directly interact with DNAs. The results suggest that numerous cellular co-solutes likely affect DNA function through these CH-pi and lone pair-pi interactions

Activation and localization of matrix metalloproteinase-2 and -9 in the skeletal muscle of the muscular dystrophy dog (CXMDJ)

<p>Abstract</p> <p>Background</p> <p>Matrix metalloproteinases (MMPs) are key regulatory molecules in the formation, remodeling and degradation of all extracellular matrix (ECM) components in both physiological and pathological processes in various tissues. The aim of this study was to examine the involvement of gelatinase MMP family members, MMP-2 and MMP-9, in dystrophin-deficient skeletal muscle. Towards this aim, we made use of the canine X-linked muscular dystrophy in Japan (CXMD_J) model, a suitable animal model for Duchenne muscular dystrophy.</p> <p>Methods</p> <p>We used surgically biopsied tibialis cranialis muscles of normal male dogs (n = 3) and CXMD_Jdogs (n = 3) at 4, 5 and 6 months of age. Muscle sections were analyzed by conventional morphological methods and <it>in situ </it>zymography to identify the localization of MMP-2 and MMP-9. MMP-2 and MMP-9 activity was examined by gelatin zymography and the levels of the respective mRNAs in addition to those of regulatory molecules, including MT1-MMP, TIMP-1, TIMP-2, and RECK, were analyzed by semi-quantitative RT-PCR.</p> <p>Results</p> <p>In CXMD_Jskeletal muscle, multiple foci of both degenerating and regenerating muscle fibers were associated with gelatinolytic MMP activity derived from MMP-2 and/or MMP-9. In CXMD_Jmuscle, MMP-9 immunoreactivity localized to degenerated fibers with inflammatory cells. Weak and disconnected immunoreactivity of basal lamina components was seen in MMP-9-immunoreactive necrotic fibers of CXMD_Jmuscle. Gelatinolytic MMP activity observed in the endomysium of groups of regenerating fibers in CXMD_Jdid not co-localize with MMP-9 immunoreactivity, suggesting that it was due to the presence of MMP-2. We observed increased activities of pro MMP-2, MMP-2 and pro MMP-9, and levels of the mRNAs encoding MMP-2, MMP-9 and the regulatory molecules, MT1-MMP, TIMP-1, TIMP-2, and RECK in the skeletal muscle of CXMD_Jdogs compared to the levels observed in normal controls.</p> <p>Conclusion</p> <p>MMP-2 and MMP-9 are likely involved in the pathology of dystrophin-deficient skeletal muscle. MMP-9 may be involved predominantly in the inflammatory process during muscle degeneration. In contrast, MMP-2, which was activated in the endomysium of groups of regenerating fibers, may be associated with ECM remodeling during muscle regeneration and fiber growth.</p

様々な物質とペプチドの複合体におけるナノ・バイオへの応用展開の検討

研究目的：近年、機能性中分子として注目されているペプチド （小タンパク質 を、さらに高機能化、多機能化させることを指向して、生体分子、無機化合物、有機小分子、高分子などと複合させた新規機能性複合体の創製を目的とする。また、これら複合体を用いたナノ分野やバイオ分野への応用展開の検討も行う 。研究概要：これまで分子単体では達成できなかったような高機能・多機能性の材料開発を目指し、無機化合物、有機小分子、高分子とペプチドを複合させた新規機能性複合体を創製する。その創製に際しては、近年、機能性中分子として注目されているペプチドを基に、ある機能を有するペプチドに、糖、核酸、酵素などの生体分子、有機合成小分子、合成高分子、無機化合物などをそれらと結合するペプチドリガンドを介して あるいは共有結合などで 複合させることで、従来以上 に 高機能化、多機能化 させた分子・複合体の創製 をねらう。さらに、本研究で作製できた複合体を用 いて、多段階触媒能を有する材料や、エレクトロニクス材料、特定疾病分子などを高度に検出できる素子、治療に役立つ薬剤などの開発につなげ、ナノ分野、バイオ分野への工学的な応用展開を検討する

Research and Design of a Routing Protocol in Large-Scale Wireless Sensor Networks

无线传感器网络，作为全球未来十大技术之一，集成了传感器技术、嵌入式计算技术、分布式信息处理和自组织网技术，可实时感知、采集、处理、传输网络分布区域内的各种信息数据，在军事国防、生物医疗、环境监测、抢险救灾、防恐反恐、危险区域远程控制等领域具有十分广阔的应用前景。 本文研究分析了无线传感器网络的已有路由协议，并针对大规模的无线传感器网络设计了一种树状路由协议，它根据节点地址信息来形成路由，从而简化了复杂繁冗的路由表查找和维护，节省了不必要的开销，提高了路由效率，实现了快速有效的数据传输。 为支持此路由协议本文提出了一种自适应动态地址分配算——ADAR（AdaptiveDynamicAddre...As one of the ten high technologies in the future, wireless sensor network, which is the integration of micro-sensors, embedded computing, modern network and Ad Hoc technologies, can apperceive, collect, process and transmit various information data within the region. It can be used in military defense, biomedical, environmental monitoring, disaster relief, counter-terrorism, remote control of haz...学位：工学硕士院系专业：信息科学与技术学院通信工程系_通信与信息系统学号：2332007115216