Determination of internucleotide (h)J(HN) couplings by the modified 2D J(NN)-correlated [(15)N, (1)H] TROSY

Recent developments in the direct observation of J couplings across hydrogen bonds in proteins and nucleic acids provide additional information for structure and function studies of these molecules by NMR spectroscopy. A J(NN)-correlated [(15)N, (1)H] TROSY experiment proposed by Pervushin et al. (Proc. Natl. Acad. Sci. USA 95, 14147-14151, 1998) can be applied to measure (h)J(HN) in smaller nucleic acids in an E.COSY manner. However, it cannot be effectively applied to large nucleic acids, such as tRNA(Trp), since one of the peaks corresponding to a fast relaxing component will be too weak to be observed in the spectra of large molecules. In this Communication, we proposed a modified J(NN)-correlated [(15)N, (1)H] TROSY experiment which enables direct measurement of (h)J(HN) in large nucleic acids.     

Application of Infrared Spectroscopy to Structure Studies of Nucleic Acids

Abstract

The purpose of this review is to show the extent to which infrared spectroscopy has been and will be useful for elucidating the molecular structures of nucleic acids.     

Nucleic acid hydration: a volumetric perspective

Hydration represents a major force governing the conformational preferences and drug binding properties of nucleic acids. Volumetric measurements have proven useful in characterizing the hydration properties of nucleic acid structures and their complexes with other molecules. In this paper, we present an overview of recent developments in the field of volumetric investigations of nucleic acids. We discuss, in particular, the volumetric properties of nucleic acids, their molecular components and analogs, conformational transitions of DNA and RNA, and drug–DNA interactions. We emphasize the importance of hydration as a major contributor to the energetics of molecular recognition. We also emphasize the need of expanding the field of volumetric characterizations of nucleic acid structures in an effort to gain further insight into the molecular origins of various nucleic acid recognition processes, including helix-to-coil and helix-to-helix conformational transitions, as well as drug–DNA and protein–DNA interactions.     

核酸-甲基青莲6B分子作用体系的研究与应用

基于核酸对甲基青莲6B的减色效应,以阳离子型染料甲基青莲6B为生色探针,研究该染料与核酸的结合反应,建立了新的核酸测定体系,研究探讨了体系的作用机理。在pH＝9.0条件下,hs-DNA,sm-DNA, ct-DNA,yeast RNA的浓度与甲基青莲6B减色效应成线性关系,响应线性范围分别为0.50～4.00 μg·mL-1,0.20～5.00 μg·mL-1,0.20～5.00 μg·mL-1,0.20～4.50 μg·mL-1,检出限(3σ/K)分别为0.082,0.037,0.038,0.041 μg·mL-1。分析核酸合成样品,回收率为93.5%～105.0%。     

Determination of Internucleotide JHN Couplings by the Modified 2D JNN-Correlated [N, H] TROSY

Recent developments in the direct observation of J couplings across hydrogen bonds in proteins and nucleic acids provide additional information for structure and function studies of these molecules by NMR spectroscopy. A J_NN-correlated [¹⁵N, ¹H] TROSY experiment proposed by Pervushin et al. (Proc. Natl. Acad. Sci. USA 95, 14147–14151, 1998) can be applied to measure ^hJ_HN in smaller nucleic acids in an E.COSY manner. However, it cannot be effectively applied to large nucleic acids, such as tRNA^Trp, since one of the peaks corresponding to a fast relaxing component will be too weak to be observed in the spectra of large molecules. In this Communication, we proposed a modified J_NN-correlated [¹⁵N, ¹H] TROSY experiment which enables direct measurement of ^hJ_HN in large nucleic acids.     

Advances in magnetofection—magnetically guided nucleic acid delivery

Magnetofection is nucleic acid delivery to cells supported and site-specifically guided by the attractive forces of magnetic fields acting on nucleic acid shuttles (vectors) which are associated with magnetic nanoparticles. Recent progress with the method confirms its general applicability with small and large nucleic acids and viruses. The method's therapeutic application as well as mechanistic studies will be discussed.     

SPECTROSCOPIC METHODS FOR CHARACTERIZATION OF NONVIRAL GENE DELIVERY SYSTEMS FROM A PHARMACEUTICAL POINT OF VIEW

Introduction of nucleic acids into mammalian and plant cells has become, in recent years, a well established procedure in basic and applied research, and the methods involved therein are now useful for the development of gene-based medicines as a new class of pharmaceuticals.

The incorporation of the gene of interest into a plasmid construct by molecular biology techniques, the complexation and condensation of nucleic acids by means of cationic agents, is the first step for enabling DNA to get into the cells of interest in a biologically active state. Monitoring this condensation procedure for scientific and quality control purposes is mostly performed by optical spectroscopy methods.

A fluorimetric, high-throughput-assay has been developed for the characterization of non-viral gene delivery systems. Protamine sulfate and cationic DOTAP liposomes were used to condense the plasmid DNA. The DNA was labeled with nucleic acid stains, and the quenching of fluorescence by the addition of condensing agents was measured. PicoGreen? was compared to a number of common nucleic acid stains (ethidium bromide, ethidium- homodimer-2, acridine orange, BOBO-1, POPO-1, TOTO-1, YOYO-1, BOBO-3, POPO-3, TOTO-3, YOYO-3, BO-PRO-1, PO-PRO-1, TO-PRO-1, YO-PRO-1, BO-PRO-3, PO-PRO-3, TO-PRO-3, YO-PRO-3), and was found to be an advantageous nucleic acid stain for the fluorimetric characterization of condensation. Stability of nucleic acid-dye complex, reproducibility and the signal-to-noise ratio make PicoGreen? a dye of choice for condensation experiments.

Condensation was quantified by the fitting of condensation curves to the “Hill equation”. Relative fluorescence intensity, Hill slope and kinetics of DNA-condensation provide the calculated values for advanced characterization of DNA-condensation.

Fluorescence spectroscopy are compared to agarose gel electrophoresis as another approach for biophysical characterization of DNA-condensation. Advantages and disadvantages of the two methods are discussed.     

解读生命密码——人类基因组计划

DNA测序技术的自动化使得人类基因组测序工作在启动10年后就已接近完成,从物理学的角度来,生物体是工作在单分子水平上的多层次综合的信息、能量和物质加工转换系统,关于生物生长发育和遗传的信息记录在线型分子——核酸的碱基序列中,每个基因是编码一个蛋白质的核酸片段,这些蛋白质分子是机器的主要零部件,首次人类基因组测序的完成,只是生命密码破译的开始,而不是结束,年轻的物理学家应积极地参与揭示生命本质的活动。     

Effect of cooling conditions on the magnetic structure of multiferroic BiFeO3 synthesized by mechanical activation

We have studied the nuclear quadrupole interactions (NQI) of the ¹⁴N, ¹⁷O and ²H nuclei in the nucleobases cytosine, adenine, guanine and thymine in the free state as well as when they are bonded to the sugar ring in DNA, simulated through a CH₃ group attached to the nucleobases. The nucleobase uracil, which replaces thymine in RNA, has also been studied. Our results show that there are substantial indirect effects of the bonding with the sugar group in the nucleic acids on the NQI parameters e ² qQ/h and η. It is hoped that measurements of these NQI parameters in DNA will be available in the future to compare with our predictions. Our results provide the conclusion that for any property dependent on the electronic structures of the nucleic acids, the effects of the bonding between the nucleobases and the nucleic acid backbones have to be included.     

Determination of Nucleic Acids Sensitized by Emulsifier OP‐micelle Using Ethyl Rhodamine B as a Resonance Light‐Scattering Probe

Abstract

The resonance light scattering (RLS) spectra of ethyl rhodamine B with nucleic acid (calf thymus DNA and herring sperm DNA) have been studied. The effective factors and the optimum conditions have been studied, and the enhanced intensity of RLS is in proportion to the concentration of nucleic acids in the range 0～5.00 µg mL^?1 for calf thymus DNA (ctDNA) and 0～3.50 µg mL^?1 for herring sperm DNA (hsDNA). The limits of detection are 3.42 and 3.14 ng mL^?1, respectively. Based on this, a RLS method for the determination of nucleic acids sensitized by emulsifier OP‐micelle was accordingly established. The binding mode concerning the interactions of ethyl rhodamine B with nucleic acids was also studied, and this method has good selectivity and high sensitivity and it has been applied to the determination of DNA in synthetic samples and real samples with satisfactory results.     

Spectroscopic Study on the Interaction of Pyronine Y with Nucleic Acids and Its Analytical Application

ABSTRACT

The interaction of pyronine Y (PY) with nucleic acids was studied by resonance Rayleigh scattering (RRS) for nucleic acid detection. The enhanced RRS intensity of nucleic acids reacted with PY was proportional to the concentration of nucleic acids in the ranges of 27.0–625 ng ml^?1 for fish sperm DNA, 39.0–500 ng ml^?1 for calf thymus DNA, and 59.0–375 ng ml^?1 for yeast RNA. The limits of determination were 0.2 ng ml^?1 for fish sperm DNA, 0.6 ng ml^?1 for calf thymus DNA, and 0.7 ng ml^?1 for yeast RNA. The method had been successfully applied to the quick determination of nucleic acids in synthetic and natural samples.     

Coassembly of Gold Nanoclusters with Nucleic Acids: Sensing,Bioimaging, and Gene Transfection

Assemblies of biopolymers and nanomaterials have become a powerful tool to build up new architectures with growing application potential. Herein, novel peptide‐stabilized fluorescent gold nanoclusters, K4‐AuNCs, are utilized as building blocks to investigate their coassembly with nucleic acids. K4‐AuNCs possess ultrasmall size (1.8 nm), red fluorescence emission, and positively charged surfaces, which are able to coassemble with DNA or RNA strands through electrostatic interactions to form pitaya‐like hybrid nanoparticles ranging from 30 to 80 nm, accompanied by an up to 3.5‐fold fluorescence enhancement. The coassembly also forms intracellularly, rendering K4‐AuNCs an attractive dye for specific in vivo nucleic acid staining, due to their higher photostability than commercial fluorescent probes such as SYTO 9. This work also demonstrates that the coassembly of K4‐AuNCs with nucleic acids can be applied to gene transfection and to build up a sensing platform to detect DNase/RNase activity or to screen their inhibitors. The new strategy greatly extends the application range of gold nanoclusters into the development of new nucleic acid detection methods and novel hybrid materials.     

荧光光谱法测定核酸的研究

研究了花菁Ⅱ与核酸的相互作用,发现在核酸的存在下,花菁Ⅱ的荧光被明显地猝灭,且猝灭程度与核酸的浓度存在良好的线性关系.     

DNA and PNA as templates for building nanoassemblies via electrostatic complexation with gold nanoparticles

Organisation of nanoparticles on structurally well-defined templates is a first step towards creating nanomachines. In this respect, nucleic acids are ideal structural templates and a variety of secondary structures realizable from DNA/RNA––e.g., duplexes, hairpins, triplexes, cruciforms, tetraplexes can be exploited to engineer nanoparticle organization at will. We have used oligonucleotides and their analogues such as phosphorothioates and peptide nucleic acids to electrostatically encapsulate cationic-capped gold nanoparticles. This article describes synthesis and characterization of DNA/PNA-gold nanoparticle composites using TEM and UV-T_m techniques. These types of assemblies may have potential for creating nanowires and lithographic circuits.     

To what extent of ion neutralization can multivalent ion distributions around RNA-like macroions be described by Poisson-Boltzmann theory?

Nucleic acids are negatively charged biomolecules, and metal ions in solutions are important to their folding structures and thermodynamics, especially multivalent ions. However, it has been suggested that the binding of multivalent ions to nucleic acids cannot be quantitatively described by the well-established Poisson-Boltzmann(PB) theory. In this work, we made extensive calculations of ion distributions around various RNA-like macroions in divalent and trivalent salt solutions by PB theory and Monte Carlo(MC) simulations. Our calculations show that PB theory appears to underestimate multivalent ion distributions around RNA-like macroions while can reliably predict monovalent ion distributions. Our extensive comparisons between PB theory and MC simulations indicate that when an RNA-like macroion gets ion neutralization beyond a "critical" value, the multivalent ion distribution around that macroion can be approximately described by PB theory.Furthermore, an empirical formula was obtained to approximately quantify the critical ion neutralization for various RNAlike macroions in multivalent salt solutions, and this empirical formula was shown to work well for various real nucleic acids including RNAs and DNAs.     

Study of the Reaction Between the Nucleic Acid and Y-BPMPHD-CTMAB Complex and Its Analytical Application

The fluorescence quenching of the Y-BPMPHD-CTMAB by nucleic acids is reported. It is considered that the Y-BPMPHD-CTMAB can form a large complex with nucleic acid through the electrostatic attraction in the pH range of 4.2-6.8. Under optimal conditions, the difference of fluorescence intensity between the system without and with nucleic acids is proportional to the concentration of nucleic acids over the range of 4.5 x 10(-8)-1.2 x 10(-5) g/mL for fsDNA and 3.2 x 10(-8)-3.0 x 10(-5) g/mL for yRNA, respectively. The detection limits are 14.0 ng/mL for fsDNA and 21.0 ng/mL for yRNA. The method is applied for the determination of nucleic acids in actual sample, and the result obtained is satisfactory.     

Tracing Life back to elements

The most ancient sequences, of proteins and nucleic acids, active in their early times, can be still recognized amongst modern sequences. That allows reconstruction of all steps of evolution of the triplet code, and of the earliest stages of evolution of genes and proteins. According to the reconstruction, the very first protein activities of living matter have been ATP binding and ATPase, responsible for energy supply. Respective modern proteins and genes are projected back to their earliest primordial forms. In particular, it is demonstrated that these two earliest activities have been encoded originally in the complementary strands of the same duplex gene.     

基于功能性核酸识别的荧光各向异性分析方法与应用

荧光各向异性方法又称荧光偏振法,基于相互作用的分子结合前后发射光退偏振的不同而实现对相互作用的研究或对目标物的检测。20世纪50年代Gregorio Weber用荧光各向异性法研究了氮磺酰氯与牛血清白蛋白和卵清白蛋白的作用,开启了该方法在生物化学研究中应用的先河。而20世纪90年代开始的功能性核酸(FNAs,包括核酸适配体和核酸酶等)的发现与合成,使基于功能性核酸的传感得到了广泛的应用。核酸适配体能够特异性识别目标分子,基于核酸识别的荧光各向异性分析方法具有高选择性、高灵敏度、高通量等优势,在研究蛋白质、核酸和小分子的相互作用中起到重要作用,然而如何提高结合前后的荧光各向异性信号变化,尤其是小分子识别前后,在基于功能性核酸识别的分析方法发展中是一个挑战。介绍了基于功能性核酸识别的荧光各向异性的方法应用于检测蛋白质、核酸及其他在生命活动中起重要作用的小分子的基本原理及设计理念。     

Spectroscopic Studies of α,γ-Disubstituted Trimethine Cyanine: New Fluorescent Dye for Nucleic Acids

Spectral properties of 3-methyl-2-3-[3-methyl-1,3-benzothiazolo-2(3H)-ylidene]-1,4-cylopentadien-1-yl-1,3-benzothiazolo-3-ium tosylate (Cyan-Cpentd) in a free state and in the complexes with nucleic acids and synthetic polynucleotides have been investigated by absorption and fluorescence spectroscopy. Significant fluorescence intensity enhancement of dye-nucleic acids complexes is observed. For the first time Cyan-Cpentd is proposed as a new probe for nucleic acid detection. Binding mechanism of Cyan-Cpentd is discussed in view of the NA-ligand interaction models.     

An Investigation of Tricarbocyanines “Stains-All” and “iso-Stains-All” as Fluorescent Nucleic Acids Probes

Luminescent properties of carbocyanine dyes Stains-All and its isomer iso-Stains-All were studied in the presence of nucleic acids. Both dyes show sufficient fluorescent intensity increase in the presence of DNA and RNA and may be used as fluorescent probes for nucleic acids (NA) detection in homogeneous assays. It was supposed that Stains-All and iso-Stains-All bind with nucleic acids through different interaction modes.