Dynamic Properties of DNA-Programmable Nanoparticle Crystallization.

PubMed

Yu, Qiuyan; Zhang, Xuena; Hu, Yi; Zhang, Zhihao; Wang, Rong

2016-08-23

The dynamics of DNA hybridization is very important in DNA-programmable nanoparticle crystallization. Here, coarse-grained molecular dynamics is utilized to explore the structural and dynamic properties of DNA hybridizations for a self-complementary DNA-directed nanoparticle self-assembly system. The hexagonal close-packed (HCP) and close-packed face-centered cubic (FCC) ordered structures are identified for the systems of different grafted DNA chains per nanoparticle, which are in good agreement with the experimental results. Most importantly, the dynamic crystallization processes of DNA hybridizations are elucidated by virtue of the mean square displacement, the percentage of hybridizations, and the lifetime of DNA bonds. The lifetime can be modeled by the DNA dehybridization, which has an exponential form. The lifetime of DNA bonds closely depends on the temperature. A suitable temperature for the DNA-nanoparticle crystallization is obtained in the work. Moreover, a too large volume fraction hinders the self-assembly process due to steric effects. This work provides some essential information for future design of nanomaterials.

Underwound DNA under Tension: Structure, Elasticity, and Sequence-Dependent Behaviors

NASA Astrophysics Data System (ADS)

Sheinin, Maxim Y.; Forth, Scott; Marko, John F.; Wang, Michelle D.

2011-09-01

DNA melting under torsion plays an important role in a wide variety of cellular processes. In the present Letter, we have investigated DNA melting at the single-molecule level using an angular optical trap. By directly measuring force, extension, torque, and angle of DNA, we determined the structural and elastic parameters of torsionally melted DNA. Our data reveal that under moderate forces, the melted DNA assumes a left-handed structure as opposed to an open bubble conformation and is highly torsionally compliant. We have also discovered that at low forces melted DNA properties are highly dependent on DNA sequence. These results provide a more comprehensive picture of the global DNA force-torque phase diagram.

Unveiling Stability Criteria of DNA-Carbon Nanotubes Constructs by Scanning Tunneling Microscopy and Computational Modeling

DOE PAGES

Kilina, Svetlana; Yarotski, Dzmitry A.; Talin, A. Alec; ...

2011-01-01

We present a combined approach that relies on computational simulations and scanning tunneling microscopy (STM) measurements to reveal morphological properties and stability criteria of carbon nanotube-DNA (CNT-DNA) constructs. Application of STM allows direct observation of very stable CNT-DNA hybrid structures with the well-defined DNA wrapping angle of 63.4 ° and a coiling period of 3.3 nm. Using force field simulations, we determine how the DNA-CNT binding energy depends on the sequence and binding geometry of a single strand DNA. This dependence allows us to quantitatively characterize the stability of a hybrid structure with an optimal π-stacking between DNA nucleotides and themore » tube surface and better interpret STM data. Our simulations clearly demonstrate the existence of a very stable DNA binding geometry for (6,5) CNT as evidenced by the presence of a well-defined minimum in the binding energy as a function of an angle between DNA strand and the nanotube chiral vector. This novel approach demonstrates the feasibility of CNT-DNA geometry studies with subnanometer resolution and paves the way towards complete characterization of the structural and electronic properties of drug-delivering systems based on DNA-CNT hybrids as a function of DNA sequence and a nanotube chirality.« less

MD simulations of papillomavirus DNA-E2 protein complexes hints at a protein structural code for DNA deformation.

PubMed

Falconi, M; Oteri, F; Eliseo, T; Cicero, D O; Desideri, A

2008-08-01

The structural dynamics of the DNA binding domains of the human papillomavirus strain 16 and the bovine papillomavirus strain 1, complexed with their DNA targets, has been investigated by modeling, molecular dynamics simulations, and nuclear magnetic resonance analysis. The simulations underline different dynamical features of the protein scaffolds and a different mechanical interaction of the two proteins with DNA. The two protein structures, although very similar, show differences in the relative mobility of secondary structure elements. Protein structural analyses, principal component analysis, and geometrical and energetic DNA analyses indicate that the two transcription factors utilize a different strategy in DNA recognition and deformation. Results show that the protein indirect DNA readout is not only addressable to the DNA molecule flexibility but it is finely tuned by the mechanical and dynamical properties of the protein scaffold involved in the interaction.

DNA materials: bridging nanotechnology and biotechnology.

PubMed

Yang, Dayong; Hartman, Mark R; Derrien, Thomas L; Hamada, Shogo; An, Duo; Yancey, Kenneth G; Cheng, Ru; Ma, Minglin; Luo, Dan

2014-06-17

CONSPECTUS: In recent decades, DNA has taken on an assortment of diverse roles, not only as the central genetic molecule in biological systems but also as a generic material for nanoscale engineering. DNA possesses many exceptional properties, including its biological function, biocompatibility, molecular recognition ability, and nanoscale controllability. Taking advantage of these unique attributes, a variety of DNA materials have been created with properties derived both from the biological functions and from the structural characteristics of DNA molecules. These novel DNA materials provide a natural bridge between nanotechnology and biotechnology, leading to far-ranging real-world applications. In this Account, we describe our work on the design and construction of DNA materials. Based on the role of DNA in the construction, we categorize DNA materials into two classes: substrate and linker. As a substrate, DNA interfaces with enzymes in biochemical reactions, making use of molecular biology's "enzymatic toolkit". For example, employing DNA as a substrate, we utilized enzymatic ligation to prepare the first bulk hydrogel made entirely of DNA. Using this DNA hydrogel as a structural scaffold, we created a protein-producing DNA hydrogel via linking plasmid DNA onto the hydrogel matrix through enzymatic ligation. Furthermore, to fully make use of the advantages of both DNA materials and polymerase chain reaction (PCR), we prepared thermostable branched DNA that could remain intact even under denaturing conditions, allowing for their use as modular primers for PCR. Moreover, via enzymatic polymerization, we have recently constructed a physical DNA hydrogel with unique internal structure and mechanical properties. As a linker, we have used DNA to interface with other functional moieties, including gold nanoparticles, clay minerals, proteins, and lipids, allowing for hybrid materials with unique properties for desired applications. For example, we recently designed a DNA-protein conjugate as a universal adapter for protein detection. We further demonstrate a diverse assortment of applications for these DNA materials including diagnostics, protein production, controlled drug release systems, the exploration of life evolution, and plasmonics. Although DNA has shown great potential as both substrate and linker in the construction of DNA materials, it is still in the initial stages of becoming a well-established and widely used material. Important challenges include the ease of design and fabrication, scaling-up, and minimizing cost. We envision that DNA materials will continue to bridge the gap between nanotechnology and biotechnology and will ultimately be employed for many real-world applications.

Characterization of the DNA binding properties of polyomavirus capsid protein

NASA Technical Reports Server (NTRS)

Chang, D.; Cai, X.; Consigli, R. A.; Spooner, B. S. (Principal Investigator)

1993-01-01

The DNA binding properties of the polyomavirus structural proteins VP1, VP2, and VP3 were studied by Southwestern analysis. The major viral structural protein VP1 and host-contributed histone proteins of polyomavirus virions were shown to exhibit DNA binding activity, but the minor capsid proteins VP2 and VP3 failed to bind DNA. The N-terminal first five amino acids (Ala-1 to Lys-5) were identified as the VP1 DNA binding domain by genetic and biochemical approaches. Wild-type VP1 expressed in Escherichia coli (RK1448) exhibited DNA binding activity, but the N-terminal truncated VP1 mutants (lacking Ala-1 to Lys-5 and Ala-1 to Cys-11) failed to bind DNA. The synthetic peptide (Ala-1 to Cys-11) was also shown to have an affinity for DNA binding. Site-directed mutagenesis of the VP1 gene showed that the point mutations at Pro-2, Lys-3, and Arg-4 on the VP1 molecule did not affect DNA binding properties but that the point mutation at Lys-5 drastically reduced DNA binding affinity. The N-terminal (Ala-1 to Lys-5) region of VP1 was found to be essential and specific for DNA binding, while the DNA appears to be non-sequence specific. The DNA binding domain and the nuclear localization signal are located in the same N-terminal region.

Structural basis for inhibition of DNA replication by aphidicolin

DOE PAGES

Baranovskiy, A. G.; Babayeva, N. D.; Suwa, Y.; ...

2014-11-27

Natural tetracyclic diterpenoid aphidicolin is a potent and specific inhibitor of B-family DNA polymerases, haltering replication and possessing a strong antimitotic activity in human cancer cell lines. Clinical trials revealed limitations of aphidicolin as an antitumor drug because of its low solubility and fast clearance from human plasma. The absence of structural information hampered the improvement of aphidicolin-like inhibitors: more than 50 modifications have been generated so far, but all have lost the inhibitory and antitumor properties. Here we report the crystal structure of the catalytic core of human DNA polymerase α (Pol α) in the ternary complex with anmore » RNA-primed DNA template and aphidicolin. The inhibitor blocks binding of dCTP by docking at the Pol α active site and by rotating the template guanine. The structure provides a plausible mechanism for the selectivity of aphidicolin incorporation opposite template guanine and explains why previous modifications of aphidicolin failed to improve its affinity for Pol α. With new structural information, aphidicolin becomes an attractive lead compound for the design of novel derivatives with enhanced inhibitory properties for B-family DNA polymerases.« less

Excited states of protonated DNA/RNA bases.

PubMed

Berdakin, Matias; Féraud, Géraldine; Dedonder-Lardeux, Claude; Jouvet, Christophe; Pino, Gustavo A

2014-06-14

The very fast relaxation of the excited states to the ground state in DNA/RNA bases is a necessary process to ensure the photostability of DNA and its rate is highly sensitive to the tautomeric form of the bases. Protonation of the bases plays a crucial role in many biochemical and mutagenic processes and it can result in alternative tautomeric structures, thus making important the knowledge of the properties of protonated DNA/RNA bases. We report here the photofragmentation spectra of the five protonated DNA/RNA bases. In most of the cases, the spectra exhibit well resolved vibrational structures, with broad bands associated with very short excited state lifetimes. The similarity between the electronic properties, e.g. excitation energy and very short excited state lifetimes for the canonical tautomers of protonated and neutral DNA bases, suggests that the former could also play an important role in the photostability mechanism of DNA.

Understanding the mechanical properties of DNA origami tiles and controlling the kinetics of their folding and unfolding reconfiguration.

PubMed

Chen, Haorong; Weng, Te-Wei; Riccitelli, Molly M; Cui, Yi; Irudayaraj, Joseph; Choi, Jong Hyun

2014-05-14

DNA origami represents a class of highly programmable macromolecules that can go through conformational changes in response to external signals. Here we show that a two-dimensional origami rectangle can be effectively folded into a short, cylindrical tube by connecting the two opposite edges through the hybridization of linker strands and that this process can be efficiently reversed via toehold-mediated strand displacement. The reconfiguration kinetics was experimentally studied as a function of incubation temperature, initial origami concentration, missing staples, and origami geometry. A kinetic model was developed by introducing the j factor to describe the reaction rates in the cyclization process. We found that the cyclization efficiency (j factor) increases sharply with temperature and depends strongly on the structural flexibility and geometry. A simple mechanical model was used to correlate the observed cyclization efficiency with origami structure details. The mechanical analysis suggests two sources of the energy barrier for DNA origami folding: overcoming global twisting and bending the structure into a circular conformation. It also provides the first semiquantitative estimation of the rigidity of DNA interhelix crossovers, an essential element in structural DNA nanotechnology. This work demonstrates efficient DNA origami reconfiguration, advances our understanding of the dynamics and mechanical properties of self-assembled DNA structures, and should be valuable to the field of DNA nanotechnology.

Characterizing DNA Star-Tile-Based Nanostructures Using a Coarse-Grained Model.

PubMed

Schreck, John S; Romano, Flavio; Zimmer, Matthew H; Louis, Ard A; Doye, Jonathan P K

2016-04-26

We use oxDNA, a coarse-grained model of DNA at the nucleotide level, to simulate large nanoprisms that are composed of multi-arm star tiles, in which the size of bulge loops that have been incorporated into the tile design is used to control the flexibility of the tiles. The oxDNA model predicts equilibrium structures for several different nanoprism designs that are in excellent agreement with the experimental structures as measured by cryoTEM. In particular we reproduce the chiral twisting of the top and bottom faces of the nanoprisms, as the bulge sizes in these structures are varied due to the greater flexibility of larger bulges. We are also able to follow how the properties of the star tiles evolve as the prisms are assembled. Individual star tiles are very flexible, but their structures become increasingly well-defined and rigid as they are incorporated into larger assemblies. oxDNA also finds that the experimentally observed prisms are more stable than their inverted counterparts, but interestingly this preference for the arms of the tiles to bend in a given direction only emerges after they are part of larger assemblies. These results show the potential for oxDNA to provide detailed structural insight as well as to predict the properties of DNA nanostructures and hence to aid rational design in DNA nanotechnology.

Mechanical design of DNA nanostructures

NASA Astrophysics Data System (ADS)

Castro, Carlos E.; Su, Hai-Jun; Marras, Alexander E.; Zhou, Lifeng; Johnson, Joshua

2015-03-01

Structural DNA nanotechnology is a rapidly emerging field that has demonstrated great potential for applications such as single molecule sensing, drug delivery, and templating molecular components. As the applications of DNA nanotechnology expand, a consideration of their mechanical behavior is becoming essential to understand how these structures will respond to physical interactions. This review considers three major avenues of recent progress in this area: (1) measuring and designing mechanical properties of DNA nanostructures, (2) designing complex nanostructures based on imposed mechanical stresses, and (3) designing and controlling structurally dynamic nanostructures. This work has laid the foundation for mechanically active nanomachines that can generate, transmit, and respond to physical cues in molecular systems.Structural DNA nanotechnology is a rapidly emerging field that has demonstrated great potential for applications such as single molecule sensing, drug delivery, and templating molecular components. As the applications of DNA nanotechnology expand, a consideration of their mechanical behavior is becoming essential to understand how these structures will respond to physical interactions. This review considers three major avenues of recent progress in this area: (1) measuring and designing mechanical properties of DNA nanostructures, (2) designing complex nanostructures based on imposed mechanical stresses, and (3) designing and controlling structurally dynamic nanostructures. This work has laid the foundation for mechanically active nanomachines that can generate, transmit, and respond to physical cues in molecular systems. Electronic supplementary information (ESI) available. See DOI: 10.1039/c4nr07153k

Deppdb--DNA electrostatic potential properties database: electrostatic properties of genome DNA.

PubMed

Osypov, Alexander A; Krutinin, Gleb G; Kamzolova, Svetlana G

2010-06-01

The electrostatic properties of genome DNA influence its interactions with different proteins, in particular, the regulation of transcription by RNA-polymerases. DEPPDB--DNA Electrostatic Potential Properties Database--was developed to hold and provide all available information on the electrostatic properties of genome DNA combined with its sequence and annotation of biological and structural properties of genome elements and whole genomes. Genomes in DEPPDB are organized on a taxonomical basis. Currently, the database contains all the completely sequenced bacterial and viral genomes according to NCBI RefSeq. General properties of the genome DNA electrostatic potential profile and principles of its formation are revealed. This potential correlates with the GC content but does not correspond to it exactly and strongly depends on both the sequence arrangement and its context (flanking regions). Analysis of the promoter regions for bacterial and viral RNA polymerases revealed a correspondence between the scale of these proteins' physical properties and electrostatic profile patterns. We also discovered a direct correlation between the potential value and the binding frequency of RNA polymerase to DNA, supporting the idea of the role of electrostatics in these interactions. This matches a pronounced tendency of the promoter regions to possess higher values of the electrostatic potential.

Unique Dynamic Properties of DNA Duplexes Containing Interstrand Crosslinks†

PubMed Central

Friedman, Joshua I.; Jiang, Yu Lin; Miller, Paul S.; Stivers, James T.

2010-01-01

Bifunctional DNA alkylating agents form a diverse assortment of covalent DNA interstrand crosslinked (ICL) structures that are potent cytotoxins. Since it is implausible that cells could possess distinct DNA repair systems for each individual ICL, it is believed that common structural and dynamic features of ICL damage are recognized, rather than specific structural characteristics of each cross-linking agent. Investigation of the structural and dynamic properties of ICLs that might be important for recognition has been complicated by heterogeneous incorporation of these lesions into DNA. To address this problem we have synthesized and characterized several homogenous ICL-DNAs containing site–specific staggered N4-cytosine-ethyl-N4-cytosine crosslinks. Staggered crosslinks were introduced in two ways: in a manner that preserves the overall structure of B-form duplex DNA, and in a manner that highly distorts the DNA structure, with the goal of understanding how structural and dynamic properties of diverse ICL duplexes might flag these sites for repair. Measurements of base pair opening dynamics in the B-form ICL duplex by 1H NMR linewidth or imino proton solvent exchange showed that the guanine base opposite to the crosslinked cytosine opened at least an order of magnitude more slowly than when in a control matched normal duplex. To a lesser degree, the B-form ICL also induced a decrease in base pair opening dynamics that extended from the site of the crosslink to adjacent base pairs. In contrast, the non-B-form ICL showed extensive conformational dynamics at the site of the cross link, which extended over the entire DNA sequence. Since DNA duplexes containing the B-form and non-B-form ICL crosslinks have both been shown to be incised when incubated in mammalian whole cell extracts, while a matched normal duplex is not, we conclude that intrinsic DNA dynamics is not a requirement for specific damage incision of these ICLs. Instead, we propose a general model where destabilized ICL-duplexes serve to energetically facilitate binding of DNA repair factors that must induce bubbles or other distortions in the duplex. However, the essential requirement for incision is an immobile Y-junction where the repair factors are stably bound at the site of the ICL, and the two DNA strands are unpaired. PMID:21174443

DNA nanotechnology and fluorescence applications.

PubMed

Schlichthaerle, Thomas; Strauss, Maximilian T; Schueder, Florian; Woehrstein, Johannes B; Jungmann, Ralf

2016-06-01

Structural DNA nanotechnology allow researchers to use the unique molecular recognition properties of DNA strands to construct nanoscale objects with almost arbitrary complexity in two and three dimensions. Abstracted as molecular breadboards, DNA nanostructures enable nanometer-precise placement of guest molecules such as proteins, fluorophores, or nanoparticles. These assemblies can be used to study biological phenomena with unprecedented control over number, spacing, and molecular identity. Here, we give a general introduction to structural DNA nanotechnology and more specifically discuss applications of DNA nanostructures in the field of fluorescence and plasmonics. Copyright © 2016 Elsevier Ltd. All rights reserved.

DNA nanotechnology based on i-motif structures.

PubMed

Dong, Yuanchen; Yang, Zhongqiang; Liu, Dongsheng

2014-06-17

CONSPECTUS: Most biological processes happen at the nanometer scale, and understanding the energy transformations and material transportation mechanisms within living organisms has proved challenging. To better understand the secrets of life, researchers have investigated artificial molecular motors and devices over the past decade because such systems can mimic certain biological processes. DNA nanotechnology based on i-motif structures is one system that has played an important role in these investigations. In this Account, we summarize recent advances in functional DNA nanotechnology based on i-motif structures. The i-motif is a DNA quadruplex that occurs as four stretches of cytosine repeat sequences form C·CH(+) base pairs, and their stabilization requires slightly acidic conditions. This unique property has produced the first DNA molecular motor driven by pH changes. The motor is reliable, and studies show that it is capable of millisecond running speeds, comparable to the speed of natural protein motors. With careful design, the output of these types of motors was combined to drive micrometer-sized cantilevers bend. Using established DNA nanostructure assembly and functionalization methods, researchers can easily integrate the motor within other DNA assembled structures and functional units, producing DNA molecular devices with new functions such as suprahydrophobic/suprahydrophilic smart surfaces that switch, intelligent nanopores triggered by pH changes, molecular logic gates, and DNA nanosprings. Recently, researchers have produced motors driven by light and electricity, which have allowed DNA motors to be integrated within silicon-based nanodevices. Moreover, some devices based on i-motif structures have proven useful for investigating processes within living cells. The pH-responsiveness of the i-motif structure also provides a way to control the stepwise assembly of DNA nanostructures. In addition, because of the stability of the i-motif, this structure can serve as the stem of one-dimensional nanowires, and a four-strand stem can provide a new basis for three-dimensional DNA structures such as pillars. By sacrificing some accuracy in assembly, we used these properties to prepare the first fast-responding pure DNA supramolecular hydrogel. This hydrogel does not swell and cannot encapsulate small molecules. These unique properties could lead to new developments in smart materials based on DNA assembly and support important applications in fields such as tissue engineering. We expect that DNA nanotechnology will continue to develop rapidly. At a fundamental level, further studies should lead to greater understanding of the energy transformation and material transportation mechanisms at the nanometer scale. In terms of applications, we expect that many of these elegant molecular devices will soon be used in vivo. These further studies could demonstrate the power of DNA nanotechnology in biology, material science, chemistry, and physics.

DNA-nanoparticle assemblies go organic: macroscopic polymeric materials with nanosized features.

PubMed

Mentovich, Elad D; Livanov, Konstantin; Prusty, Deepak K; Sowwan, Mukules; Richter, Shachar

2012-05-30

One of the goals in the field of structural DNA nanotechnology is the use of DNA to build up 2- and 3-D nanostructures. The research in this field is motivated by the remarkable structural features of DNA as well as by its unique and reversible recognition properties. Nucleic acids can be used alone as the skeleton of a broad range of periodic nanopatterns and nanoobjects and in addition, DNA can serve as a linker or template to form DNA-hybrid structures with other materials. This approach can be used for the development of new detection strategies as well as nanoelectronic structures and devices. Here we present a new method for the generation of unprecedented all-organic conjugated-polymer nanoparticle networks guided by DNA, based on a hierarchical self-assembly process. First, microphase separation of amphiphilic block copolymers induced the formation of spherical nanoobjects. As a second ordering concept, DNA base pairing has been employed for the controlled spatial definition of the conjugated-polymer particles within the bulk material. These networks offer the flexibility and the diversity of soft polymeric materials. Thus, simple chemical methodologies could be applied in order to tune the network's electrical, optical and mechanical properties. One- two- and three-dimensional networks have been successfully formed. Common to all morphologies is the integrity of the micelles consisting of DNA block copolymer (DBC), which creates an all-organic engineered network.

DEPPDB - DNA electrostatic potential properties database. Electrostatic properties of genome DNA elements.

PubMed

Osypov, Alexander A; Krutinin, Gleb G; Krutinina, Eugenia A; Kamzolova, Svetlana G

2012-04-01

Electrostatic properties of genome DNA are important to its interactions with different proteins, in particular, related to transcription. DEPPDB - DNA Electrostatic Potential (and other Physical) Properties Database - provides information on the electrostatic and other physical properties of genome DNA combined with its sequence and annotation of biological and structural properties of genomes and their elements. Genomes are organized on taxonomical basis, supporting comparative and evolutionary studies. Currently, DEPPDB contains all completely sequenced bacterial, viral, mitochondrial, and plastids genomes according to the NCBI RefSeq, and some model eukaryotic genomes. Data for promoters, regulation sites, binding proteins, etc., are incorporated from established DBs and literature. The database is complemented by analytical tools. User sequences calculations are available. Case studies discovered electrostatics complementing DNA bending in E.coli plasmid BNT2 promoter functioning, possibly affecting host-environment metabolic switch. Transcription factors binding sites gravitate to high potential regions, confirming the electrostatics universal importance in protein-DNA interactions beyond the classical promoter-RNA polymerase recognition and regulation. Other genome elements, such as terminators, also show electrostatic peculiarities. Most intriguing are gene starts, exhibiting taxonomic correlations. The necessity of the genome electrostatic properties studies is discussed.

Pharmacologic Effects on Mitochondrial Function

ERIC Educational Resources Information Center

Cohen, Bruce H.

2010-01-01

The vast majority of energy necessary for cellular function is produced in mitochondria. Free-radical production and apoptosis are other critical mitochondrial functions. The complex structure, electrochemical properties of the inner mitochondrial membrane (IMM), and genetic control from both mitochondrial DNA (mtDNA) and nuclear DNA (nDNA) are…

Raman spectroscopic study of plasma-treated salmon DNA

NASA Astrophysics Data System (ADS)

Joon Lee, Geon; Kwon, Young-Wan; Hee Kim, Yong; Ha Choi, Eun

2013-01-01

In this research, we studied the effect of plasma treatment on the optical/structural properties of the deoxyribonucleic acid (DNA) extracted from salmon sperm. DNA-cetyltrimethylammonium (CTMA) films were obtained by complexation of DNA with CTMA. Circular dichroism (CD) and Raman spectra indicated that DNA retained its double helical structure in the solid film. The Raman spectra exhibited several vibration modes corresponding to the nuclear bases and the deoxyribose-phosphate backbones of the DNA, as well as the alkylchains of CTMA. Dielectric-barrier-discharge (DBD) plasma treatment induced structural modification and damage to the DNA, as observed by changes in the ultraviolet-visible absorption, CD, and Raman spectra. The optical emission spectra of the DBD plasma confirmed that DNA modification was induced by plasma ions such as reactive oxygen species and reactive nitrogen species.

Molecular engineering of chiral colloidal liquid crystals using DNA origami

NASA Astrophysics Data System (ADS)

Siavashpouri, Mahsa; Wachauf, Christian H.; Zakhary, Mark J.; Praetorius, Florian; Dietz, Hendrik; Dogic, Zvonimir

2017-08-01

Establishing precise control over the shape and the interactions of the microscopic building blocks is essential for design of macroscopic soft materials with novel structural, optical and mechanical properties. Here, we demonstrate robust assembly of DNA origami filaments into cholesteric liquid crystals, one-dimensional supramolecular twisted ribbons and two-dimensional colloidal membranes. The exquisite control afforded by the DNA origami technology establishes a quantitative relationship between the microscopic filament structure and the macroscopic cholesteric pitch. Furthermore, it also enables robust assembly of one-dimensional twisted ribbons, which behave as effective supramolecular polymers whose structure and elastic properties can be precisely tuned by controlling the geometry of the elemental building blocks. Our results demonstrate the potential synergy between DNA origami technology and colloidal science, in which the former allows for rapid and robust synthesis of complex particles, and the latter can be used to assemble such particles into bulk materials.

Molecular engineering of chiral colloidal liquid crystals using DNA origami.

PubMed

Siavashpouri, Mahsa; Wachauf, Christian H; Zakhary, Mark J; Praetorius, Florian; Dietz, Hendrik; Dogic, Zvonimir

2017-08-01

Establishing precise control over the shape and the interactions of the microscopic building blocks is essential for design of macroscopic soft materials with novel structural, optical and mechanical properties. Here, we demonstrate robust assembly of DNA origami filaments into cholesteric liquid crystals, one-dimensional supramolecular twisted ribbons and two-dimensional colloidal membranes. The exquisite control afforded by the DNA origami technology establishes a quantitative relationship between the microscopic filament structure and the macroscopic cholesteric pitch. Furthermore, it also enables robust assembly of one-dimensional twisted ribbons, which behave as effective supramolecular polymers whose structure and elastic properties can be precisely tuned by controlling the geometry of the elemental building blocks. Our results demonstrate the potential synergy between DNA origami technology and colloidal science, in which the former allows for rapid and robust synthesis of complex particles, and the latter can be used to assemble such particles into bulk materials.

Nonlinear optical and G-Quadruplex DNA stabilization properties of novel mixed ligand copper(II) complexes and coordination polymers: Synthesis, structural characterization and computational studies

NASA Astrophysics Data System (ADS)

Rajasekhar, Bathula; Bodavarapu, Navya; Sridevi, M.; Thamizhselvi, G.; RizhaNazar, K.; Padmanaban, R.; Swu, Toka

2018-03-01

The present study reports the synthesis and evaluation of nonlinear optical property and G-Quadruplex DNA Stabilization of five novel copper(II) mixed ligand complexes. They were synthesized from copper(II) salt, 2,5- and 2,3- pyridinedicarboxylic acid, diethylenetriamine and amide based ligand (AL). The crystal structure of these complexes were determined through X-ray diffraction and supported by ESI-MAS, NMR, UV-Vis and FT-IR spectroscopic methods. Their nonlinear optical property was studied using Gaussian09 computer program. For structural optimization and nonlinear optical property, density functional theory (DFT) based B3LYP method was used with LANL2DZ basis set for metal ion and 6-31G∗ for C,H,N,O and Cl atoms. The present work reveals that pre-polarized Complex-2 showed higher β value (29.59 × 10-30e.s.u) as compared to that of neutral complex-1 (β = 0.276 × 10-30e.s.u.) which may be due to greater advantage of polarizability. Complex-2 is expected to be a potential material for optoelectronic and photonic technologies. Docking studies using AutodockVina revealed that complex-2 has higher binding energy for both G-Quadruplex DNA (-8.7 kcal/mol) and duplex DNA (-10.1 kcal/mol). It was also observed that structure plays an important role in binding efficiency.

Decoding the conformation-linked functional properties of nucleic acids by the use of computational tools.

PubMed

Iacovelli, Federico; Falconi, Mattia

2015-09-01

DNA and RNA are large and flexible polymers selected by nature to transmit information. The most common DNA three-dimensional structure is represented by the double helix, but this biopolymer is extremely flexible and polymorphic, and can easily change its conformation to adapt to different interactions and purposes. DNA can also adopt singular topologies, giving rise, for instance, to supercoils, formed because of the limited free rotation of the DNA domain flanking a replication or transcription complex. Our understanding of the importance of these unusual or transient structures is growing, as recent studies of DNA topology, supercoiling, knotting and linking have shown that the geometric changes can drive, or strongly influence, the interactions between protein and DNA, so altering its own metabolism. On the other hand, the unique self-recognition properties of DNA, determined by the strict Watson-Crick rules of base pairing, make this material ideal for the creation of self-assembling, predesigned nanostructures. The construction of such structures is one of the main focuses of the thriving area of DNA nanotechnology, where several assembly strategies have been employed to build increasingly complex DNA nanostructures. DNA nanodevices can have direct applications in biomedicine, but also in the materials science field, requiring the immersion of DNA in an environment far from the physiological one. Crucial help in the understanding and planning of natural and artificial nanostructures is given by modern computer simulation techniques, which are able to provide a reliable structural and dynamic description of nucleic acids. © 2015 FEBS.

Optical Voltage Sensing Using DNA Origami

PubMed Central

2018-01-01

We explore the potential of DNA nanotechnology for developing novel optical voltage sensing nanodevices that convert a local change of electric potential into optical signals. As a proof-of-concept of the sensing mechanism, we assembled voltage responsive DNA origami structures labeled with a single pair of FRET dyes. The DNA structures were reversibly immobilized on a nanocapillary tip and underwent controlled structural changes upon application of an electric field. The applied field was monitored through a change in FRET efficiency. By exchanging the position of a single dye, we could tune the voltage sensitivity of our DNA origami structure, demonstrating the flexibility and versatility of our approach. The experimental studies were complemented by coarse-grained simulations that characterized voltage-dependent elastic deformation of the DNA nanostructures and the associated change in the distance between the FRET pair. Our work opens a novel pathway for determining the mechanical properties of DNA origami structures and highlights potential applications of dynamic DNA nanostructures as voltage sensors. PMID:29430924

DNA damage may drive nucleosomal reorganization to facilitate damage detection

NASA Astrophysics Data System (ADS)

LeGresley, Sarah E.; Wilt, Jamie; Antonik, Matthew

2014-03-01

One issue in genome maintenance is how DNA repair proteins find lesions at rates that seem to exceed diffusion-limited search rates. We propose a phenomenon where DNA damage induces nucleosomal rearrangements which move lesions to potential rendezvous points in the chromatin structure. These rendezvous points are the dyad and the linker DNA between histones, positions in the chromatin which are more likely to be accessible by repair proteins engaged in a random search. The feasibility of this mechanism is tested by considering the statistical mechanics of DNA containing a single lesion wrapped onto the nucleosome. We consider lesions which make the DNA either more flexible or more rigid by modeling the lesion as either a decrease or an increase in the bending energy. We include this energy in a partition function model of nucleosome breathing. Our results indicate that the steady state for a breathing nucleosome will most likely position the lesion at the dyad or in the linker, depending on the energy of the lesion. A role for DNA binding proteins and chromatin remodelers is suggested based on their ability to alter the mechanical properties of the DNA and DNA-histone binding, respectively. We speculate that these positions around the nucleosome potentially serve as rendezvous points where DNA lesions may be encountered by repair proteins which may be sterically hindered from searching the rest of the nucleosomal DNA. The strength of the repositioning is strongly dependent on the structural details of the DNA lesion and the wrapping and breathing of the nucleosome. A more sophisticated evaluation of this proposed mechanism will require detailed information about breathing dynamics, the structure of partially wrapped nucleosomes, and the structural properties of damaged DNA.

DNA-nanoparticle assemblies go organic: Macroscopic polymeric materials with nanosized features

PubMed Central

2012-01-01

Background One of the goals in the field of structural DNA nanotechnology is the use of DNA to build up 2- and 3-D nanostructures. The research in this field is motivated by the remarkable structural features of DNA as well as by its unique and reversible recognition properties. Nucleic acids can be used alone as the skeleton of a broad range of periodic nanopatterns and nanoobjects and in addition, DNA can serve as a linker or template to form DNA-hybrid structures with other materials. This approach can be used for the development of new detection strategies as well as nanoelectronic structures and devices. Method Here we present a new method for the generation of unprecedented all-organic conjugated-polymer nanoparticle networks guided by DNA, based on a hierarchical self-assembly process. First, microphase separation of amphiphilic block copolymers induced the formation of spherical nanoobjects. As a second ordering concept, DNA base pairing has been employed for the controlled spatial definition of the conjugated-polymer particles within the bulk material. These networks offer the flexibility and the diversity of soft polymeric materials. Thus, simple chemical methodologies could be applied in order to tune the network's electrical, optical and mechanical properties. Results and conclusions One- two- and three-dimensional networks have been successfully formed. Common to all morphologies is the integrity of the micelles consisting of DNA block copolymer (DBC), which creates an all-organic engineered network. PMID:22646980

Biophysical and electrochemical properties of Self-assembled noncovalent SWNT/DNA hybrid and electroactive nanostructure

NASA Astrophysics Data System (ADS)

Mirzapoor, Aboulfazl; Ranjbar, Bijan

2017-09-01

DNA self-assembled hybrid nanostructures are widely used in recent research in nanobiotechnology. Combination of DNA with carbon based nanoparticles such as single-walled carbon nanotube (SWNT), multi-walled carbon nanotube (MWNT) and carbon quantum dot were applied in important biological applications. Many examples of biosensors, nanowires and nanoelectronic devices, nanomachine and drug delivery systems are fabricated by these hybrid nanostructures. In this study, a new hybrid nanostructure has been fabricated by noncovalent interactions between single or double stranded DNA and SWNT nanoparticles and biophysical properties of these structures were studied comparatively. Biophysical properties of hybrid nanostructures studied by circular dichroism, UV-vis and fluorescence spectroscopy techniques. Also, electrochemical properties studied by cyclic voltammetry, linear sweep voltammetry, square wave voltammetry, choronoamperometry and impedance spectroscopy (EIS). Results revealed that the biophysical and electrochemical properties of SWNT/DNA hybrid nanostructures were different compare to ss-DNA, ds-DNA and SWNT singly. Circular dichroism results showed that ss-DNA wrapped around the nanotubes through π-π stacking interactions. The results indicated that after adding SWNT to ss-DNA and ds-DNA intensity of CD and UV-vis spectrum peaks were decreased. Electrochemical experiments indicated that the modification of single-walled carbon nanotubes by ss-DNA improves the electron transfer rate of hybrid nanostructures. It was demonstrated SWNT/DNA hybrid nanostructures should be a good electroactive nanostructure that can be used for electrochemical detection or sensing.

Spectroelectrochemical insights into structural and redox properties of immobilized endonuclease III and its catalytically inactive mutant

NASA Astrophysics Data System (ADS)

Moe, Elin; Rollo, Filipe; Silveira, Célia M.; Sezer, Murat; Hildebrandt, Peter; Todorovic, Smilja

2018-01-01

Endonuclease III is a Fe-S containing bifunctional DNA glycosylase which is involved in the repair of oxidation damaged DNA. Here we employ surface enhanced IR spectroelectrochemistry and electrochemistry to study the enzyme from the highly radiation- and desiccation-resistant bacterium Deinococcus radiodurans (DrEndoIII2). The experiments are designed to shed more light onto specific parameters that are currently proposed to govern damage search and recognition by endonucleases III. We demonstrate that electrostatic interactions required for the redox activation of DrEndoIII2 may result in high electric fields that alter its structural and thermodynamic properties. Analysis of inactive DrEndoIII2 (K132A/D150A double mutant) interacting with undamaged DNA, and the active enzyme interacting with damaged DNA also indicate that the electron transfer is modulated by subtle differences in the protein-DNA complex.

Spectroelectrochemical insights into structural and redox properties of immobilized endonuclease III and its catalytically inactive mutant.

PubMed

Moe, Elin; Rollo, Filipe; Silveira, Célia M; Sezer, Murat; Hildebrandt, Peter; Todorovic, Smilja

2018-01-05

Endonuclease III is a Fe-S containing bifunctional DNA glycosylase which is involved in the repair of oxidation damaged DNA. Here we employ surface enhanced IR spectroelectrochemistry and electrochemistry to study the enzyme from the highly radiation- and desiccation-resistant bacterium Deinococcus radiodurans (DrEndoIII 2 ). The experiments are designed to shed more light onto specific parameters that are currently proposed to govern damage search and recognition by endonucleases III. We demonstrate that electrostatic interactions required for the redox activation of DrEndoIII 2 may result in high electric fields that alter its structural and thermodynamic properties. Analysis of inactive DrEndoIII 2 (K132A/D150A double mutant) interacting with undamaged DNA, and the active enzyme interacting with damaged DNA also indicate that the electron transfer is modulated by subtle differences in the protein-DNA complex. Copyright © 2017 Elsevier B.V. All rights reserved.

Recent development in deciphering the structure of luminescent silver nanodots

NASA Astrophysics Data System (ADS)

Choi, Sungmoon; Yu, Junhua

2017-05-01

Matrix-stabilized silver clusters and stable luminescent few-atom silver clusters, referred to as silver nanodots, show notable difference in their photophysical properties. We present recent research on deciphering the nature of silver clusters and nanodots and understanding the factors that lead to variations in luminescent mechanisms. Due to their relatively simple structure, the matrix-stabilized clusters have been well studied. However, the single-stranded DNA (ssDNA)-stabilized silver nanodots that show the most diverse emission wavelengths and the best photophysical properties remain mysterious species. It is clear that their photophysical properties highly depend on their protection scaffolds. Analyses from combinations of high-performance liquid chromatography, inductively coupled plasma-atomic emission spectroscopy, electrophoresis, and mass spectrometry indicate that about 10 to 20 silver atoms form emissive complexes with ssDNA. However, it is possible that not all of the silver atoms in the complex form effective emission centers. Investigation of the nanodot structure will help us understand why luminescent silver nanodots are stable in aqueous solution and how to further improve their chemical and photophysical properties.

Analysis of DNA binding by human factor xeroderma pigmentosum complementation group A (XPA) provides insight into its interactions with nucleotide excision repair substrates.

PubMed

Sugitani, Norie; Voehler, Markus W; Roh, Michelle S; Topolska-Woś, Agnieszka M; Chazin, Walter J

2017-10-13

Xeroderma pigmentosum (XP) complementation group A (XPA) is an essential scaffolding protein in the multiprotein nucleotide excision repair (NER) machinery. The interaction of XPA with DNA is a core function of this protein; a number of mutations in the DNA-binding domain (DBD) are associated with XP disease. Although structures of the central globular domain of human XPA and data on binding of DNA substrates have been reported, the structural basis for XPA's DNA-binding activity remains unknown. X-ray crystal structures of the central globular domain of yeast XPA (Rad14) with lesion-containing DNA duplexes have provided valuable insights, but the DNA substrates used for this study do not correspond to the substrates of XPA as it functions within the NER machinery. To better understand the DNA-binding activity of human XPA in NER, we used NMR to investigate the interaction of its DBD with a range of DNA substrates. We found that XPA binds different single-stranded/double-stranded junction DNA substrates with a common surface. Comparisons of our NMR-based mapping of binding residues with the previously reported Rad14-DNA crystal structures revealed similarities and differences in substrate binding between XPA and Rad14. This includes direct evidence for DNA contacts to the residues extending C-terminally from the globular core, which are lacking in the Rad14 construct. Moreover, mutation of the XPA residue corresponding to Phe-262 in Rad14, previously reported as being critical for DNA binding, had only a moderate effect on the DNA-binding activity of XPA. The DNA-binding properties of several disease-associated mutations in the DBD were investigated. These results suggest that for XPA mutants exhibiting altered DNA-binding properties, a correlation exists between the extent of reduction in DNA-binding affinity and the severity of symptoms in XP patients. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Measurement of changes in impedance of DNA nanowires due to radiation induced structural damage. A novel approach for a DNA-based radiosensitive device

NASA Astrophysics Data System (ADS)

Heimbach, Florian; Arndt, Alexander; Nettelbeck, Heidi; Langner, Frank; Giesen, Ulrich; Rabus, Hans; Sellner, Stefan; Toppari, Jussi; Shen, Boxuan; Baek, Woon Yong

2017-08-01

The ability of DNA to conduct electric current has been the topic of numerous investigations over the past few decades. Those investigations indicate that this ability is dependent on the molecular structure of the DNA. Radiation-induced damages, which lead to an alteration of the molecular structure, should therefore change the electrical impedance of a DNA molecule. In this paper, the damage due to ionising radiation is shown to have a direct effect on the electrical transport properties of DNA. Impedance measurements of DNA samples were carried out by an AC impedance spectrometer before, during and after irradiation. The samples comprised of DNA segments, which were immobilized between gold electrodes with a gap of 12 μm. The impedance of all DNA samples exhibited rising capacitive behaviour with increasing absorbed dose.

Characterization of the geometry and topology of DNA pictured as a discrete collection of atoms

PubMed Central

Olson, Wilma K.

2014-01-01

The structural and physical properties of DNA are closely related to its geometry and topology. The classical mathematical treatment of DNA geometry and topology in terms of ideal smooth space curves was not designed to characterize the spatial arrangements of atoms found in high-resolution and simulated double-helical structures. We present here new and rigorous numerical methods for the rapid and accurate assessment of the geometry and topology of double-helical DNA structures in terms of the constituent atoms. These methods are well designed for large DNA datasets obtained in detailed numerical simulations or determined experimentally at high-resolution. We illustrate the usefulness of our methodology by applying it to the analysis of three canonical double-helical DNA chains, a 65-bp minicircle obtained in recent molecular dynamics simulations, and a crystallographic array of protein-bound DNA duplexes. Although we focus on fully base-paired DNA structures, our methods can be extended to treat the geometry and topology of melted DNA structures as well as to characterize the folding of arbitrary molecules such as RNA and cyclic peptides. PMID:24791158

Presynaptic Filament Dynamics in Homologous Recombination and DNA Repair

PubMed Central

Liu, Jie; Ehmsen, Kirk T.; Heyer, Wolf-Dietrich; Morrical, Scott W.

2014-01-01

Homologous Recombination (HR) is an essential genome stability mechanism used for high-fidelity repair of DNA double-strand breaks and for the recovery of stalled or collapsed DNA replication forks. The crucial homology search and DNA strand exchange steps of HR are catalyzed by presynaptic filaments—helical filaments of a recombinase enzyme bound to single-stranded DNA. Presynaptic filaments are fundamentally dynamic structures, the assembly, catalytic turnover, and disassembly of which must be closely coordinated with other elements of the DNA recombination, repair, and replication machinery in order for genome maintenance functions to be effective. Here, we review the major dynamic elements controlling the assembly, activity, and disassembly of presynaptic filaments: some intrinsic such as recombinase ATP binding and hydrolytic activities, others extrinsic such as ssDNA-binding proteins, mediator proteins, and DNA motor proteins. We examine dynamic behavior on multiple levels, including atomic- and filament-level structural changes associated with ATP binding and hydrolysis as evidenced in crystal structures, as well as subunit binding and dissociation events driven by intrinsic and extrinsic factors. We examine the biochemical properties of recombination proteins from four model systems (T4 phage, E. coli, S. cerevisiae, and H. sapiens), demonstrating how their properties are tailored for the context-specific requirements in these diverse species. We propose that the presynaptic filament has evolved to rely on multiple external factors for increased multi-level regulation of HR processes in genomes with greater structural and sequence complexity. PMID:21599536

Assembly and structural analysis of a covalently closed nano-scale DNA cage

PubMed Central

Andersen, Felicie F.; Knudsen, Bjarne; Oliveira, Cristiano Luis Pinto; Frøhlich, Rikke F.; Krüger, Dinna; Bungert, Jörg; Agbandje-McKenna, Mavis; McKenna, Robert; Juul, Sissel; Veigaard, Christopher; Koch, Jørn; Rubinstein, John L.; Guldbrandtsen, Bernt; Hede, Marianne S.; Karlsson, Göran; Andersen, Anni H.; Pedersen, Jan Skov; Knudsen, Birgitta R.

2008-01-01

The inherent properties of DNA as a stable polymer with unique affinity for partner molecules determined by the specific Watson–Crick base pairing makes it an ideal component in self-assembling structures. This has been exploited for decades in the design of a variety of artificial substrates for investigations of DNA-interacting enzymes. More recently, strategies for synthesis of more complex two-dimensional (2D) and 3D DNA structures have emerged. However, the building of such structures is still in progress and more experiences from different research groups and different fields of expertise are necessary before complex DNA structures can be routinely designed for the use in basal science and/or biotechnology. Here we present the design, construction and structural analysis of a covalently closed and stable 3D DNA structure with the connectivity of an octahedron, as defined by the double-stranded DNA helices that assembles from eight oligonucleotides with a yield of ∼30%. As demonstrated by Small Angle X-ray Scattering and cryo-Transmission Electron Microscopy analyses the eight-stranded DNA structure has a central cavity larger than the apertures in the surrounding DNA lattice and can be described as a nano-scale DNA cage, Hence, in theory it could hold proteins or other bio-molecules to enable their investigation in certain harmful environments or even allow their organization into higher order structures. PMID:18096620

Multiple conformations of the cytidine repressor DNA-binding domain coalesce to one upon recognition of a specific DNA surface.

PubMed

Moody, Colleen L; Tretyachenko-Ladokhina, Vira; Laue, Thomas M; Senear, Donald F; Cocco, Melanie J

2011-08-09

The cytidine repressor (CytR) is a member of the LacR family of bacterial repressors with distinct functional features. The Escherichia coli CytR regulon comprises nine operons whose palindromic operators vary in both sequence and, most significantly, spacing between the recognition half-sites. This suggests a strong likelihood that protein folding would be coupled to DNA binding as a mechanism to accommodate the variety of different operator architectures to which CytR is targeted. Such coupling is a common feature of sequence-specific DNA-binding proteins, including the LacR family repressors; however, there are no significant structural rearrangements upon DNA binding within the three-helix DNA-binding domains (DBDs) studied to date. We used nuclear magnetic resonance (NMR) spectroscopy to characterize the CytR DBD free in solution and to determine the high-resolution structure of a CytR DBD monomer bound specifically to one DNA half-site of the uridine phosphorylase (udp) operator. We find that the free DBD populates multiple distinct conformations distinguished by up to four sets of NMR peaks per residue. This structural heterogeneity is previously unknown in the LacR family. These stable structures coalesce into a single, more stable udp-bound form that features a three-helix bundle containing a canonical helix-turn-helix motif. However, this structure differs from all other LacR family members whose structures are known with regard to the packing of the helices and consequently their relative orientations. Aspects of CytR activity are unique among repressors; we identify here structural properties that are also distinct and that might underlie the different functional properties. © 2011 American Chemical Society

Structural DNA Nanotechnology: Artificial Nanostructures for Biomedical Research.

PubMed

Ke, Yonggang; Castro, Carlos; Choi, Jong Hyun

2018-06-04

Structural DNA nanotechnology utilizes synthetic or biologic DNA as designer molecules for the self-assembly of artificial nanostructures. The field is founded upon the specific interactions between DNA molecules, known as Watson-Crick base pairing. After decades of active pursuit, DNA has demonstrated unprecedented versatility in constructing artificial nanostructures with significant complexity and programmability. The nanostructures could be either static, with well-controlled physicochemical properties, or dynamic, with the ability to reconfigure upon external stimuli. Researchers have devoted considerable effort to exploring the usability of DNA nanostructures in biomedical research. We review the basic design methods for fabricating both static and dynamic DNA nanostructures, along with their biomedical applications in fields such as biosensing, bioimaging, and drug delivery.

Three-dimensional structural dynamics and fluctuations of DNA-nanogold conjugates by individual-particle electron tomography

NASA Astrophysics Data System (ADS)

Zhang, Lei; Lei, Dongsheng; Smith, Jessica M.; Zhang, Meng; Tong, Huimin; Zhang, Xing; Lu, Zhuoyang; Liu, Jiankang; Alivisatos, A. Paul; Ren, Gang

2016-03-01

DNA base pairing has been used for many years to direct the arrangement of inorganic nanocrystals into small groupings and arrays with tailored optical and electrical properties. The control of DNA-mediated assembly depends crucially on a better understanding of three-dimensional structure of DNA-nanocrystal-hybridized building blocks. Existing techniques do not allow for structural determination of these flexible and heterogeneous samples. Here we report cryo-electron microscopy and negative-staining electron tomography approaches to image, and three-dimensionally reconstruct a single DNA-nanogold conjugate, an 84-bp double-stranded DNA with two 5-nm nanogold particles for potential substrates in plasmon-coupling experiments. By individual-particle electron tomography reconstruction, we obtain 14 density maps at ~2-nm resolution. Using these maps as constraints, we derive 14 conformations of dsDNA by molecular dynamics simulations. The conformational variation is consistent with that from liquid solution, suggesting that individual-particle electron tomography could be an expected approach to study DNA-assembling and flexible protein structure and dynamics.

Mechanical design of DNA nanostructures.

PubMed

Castro, Carlos E; Su, Hai-Jun; Marras, Alexander E; Zhou, Lifeng; Johnson, Joshua

2015-04-14

Structural DNA nanotechnology is a rapidly emerging field that has demonstrated great potential for applications such as single molecule sensing, drug delivery, and templating molecular components. As the applications of DNA nanotechnology expand, a consideration of their mechanical behavior is becoming essential to understand how these structures will respond to physical interactions. This review considers three major avenues of recent progress in this area: (1) measuring and designing mechanical properties of DNA nanostructures, (2) designing complex nanostructures based on imposed mechanical stresses, and (3) designing and controlling structurally dynamic nanostructures. This work has laid the foundation for mechanically active nanomachines that can generate, transmit, and respond to physical cues in molecular systems.

Using Atomic Force Microscopy to Characterize the Conformational Properties of Proteins and Protein-DNA Complexes That Carry Out DNA Repair.

PubMed

LeBlanc, Sharonda; Wilkins, Hunter; Li, Zimeng; Kaur, Parminder; Wang, Hong; Erie, Dorothy A

2017-01-01

Atomic force microscopy (AFM) is a scanning probe technique that allows visualization of single biomolecules and complexes deposited on a surface with nanometer resolution. AFM is a powerful tool for characterizing protein-protein and protein-DNA interactions. It can be used to capture snapshots of protein-DNA solution dynamics, which in turn, enables the characterization of the conformational properties of transient protein-protein and protein-DNA interactions. With AFM, it is possible to determine the stoichiometries and binding affinities of protein-protein and protein-DNA associations, the specificity of proteins binding to specific sites on DNA, and the conformations of the complexes. We describe methods to prepare and deposit samples, including surface treatments for optimal depositions, and how to quantitatively analyze images. We also discuss a new electrostatic force imaging technique called DREEM, which allows the visualization of the path of DNA within proteins in protein-DNA complexes. Collectively, these methods facilitate the development of comprehensive models of DNA repair and provide a broader understanding of all protein-protein and protein-nucleic acid interactions. The structural details gleaned from analysis of AFM images coupled with biochemistry provide vital information toward establishing the structure-function relationships that govern DNA repair processes. © 2017 Elsevier Inc. All rights reserved.

Controlling Non-Equilibrium Structure Formation on the Nanoscale.

PubMed

Buchmann, Benedikt; Hecht, Fabian Manfred; Pernpeintner, Carla; Lohmueller, Theobald; Bausch, Andreas R

2017-12-06

Controlling the structure formation of gold nanoparticle aggregates is a promising approach towards novel applications in many fields, ranging from (bio)sensing to (bio)imaging to medical diagnostics and therapeutics. To steer structure formation, the DNA-DNA interactions of DNA strands that are coated on the surface of the particles have become a valuable tool to achieve precise control over the interparticle potentials. In equilibrium approaches, this technique is commonly used to study particle crystallization and ligand binding. However, regulating the structural growth processes from the nano- to the micro- and mesoscale remains elusive. Here, we show that the non-equilibrium structure formation of gold nanoparticles can be stirred in a binary heterocoagulation process to generate nanoparticle clusters of different sizes. The gold nanoparticles are coated with sticky single stranded DNA and mixed at different stoichiometries and sizes. This not only allows for structural control but also yields access to the optical properties of the nanoparticle suspensions. As a result, we were able to reliably control the kinetic structure formation process to produce cluster sizes between tens of nanometers up to micrometers. Consequently, the intricate optical properties of the gold nanoparticles could be utilized to control the maximum of the nanoparticle suspension extinction spectra between 525 nm and 600 nm. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

The structure and DNA-binding properties of Mgm101 from a yeast with a linear mitochondrial genome

PubMed Central

Pevala, Vladimír; Truban, Dominika; Bauer, Jacob A.; Košťan, Július; Kunová, Nina; Bellová, Jana; Brandstetter, Marlene; Marini, Victoria; Krejčí, Lumír; Tomáška, Ľubomír; Nosek, Jozef; Kutejová, Eva

2016-01-01

To study the mechanisms involved in the maintenance of a linear mitochondrial genome we investigated the biochemical properties of the recombination protein Mgm101 from Candida parapsilosis. We show that CpMgm101 complements defects associated with the Saccharomyces cerevisiae mgm101–1ts mutation and that it is present in both the nucleus and mitochondrial nucleoids of C. parapsilosis. Unlike its S. cerevisiae counterpart, CpMgm101 is associated with the entire nucleoid population and is able to bind to a broad range of DNA substrates in a non-sequence specific manner. CpMgm101 is also able to catalyze strand annealing and D-loop formation. CpMgm101 forms a roughly C-shaped trimer in solution according to SAXS. Electron microscopy of a complex of CpMgm101 with a model mitochondrial telomere revealed homogeneous, ring-shaped structures at the telomeric single-stranded overhangs. The DNA-binding properties of CpMgm101, together with its DNA recombination properties, suggest that it can play a number of possible roles in the replication of the mitochondrial genome and the maintenance of its telomeres. PMID:26743001

Programmable Self-Assembly of DNA-Dendrimer and DNA-Fullerene Nanostructures

DTIC Science & Technology

2004-10-01

separated by high pressure liquid chromatography ( HPLC ). The resulting material was analytically pure (99%) and monodisperse. Hybridization...bacterial and viral recognition, and gene expression analysis . These major accomplishments have been disseminated by various applications including 16...designing DNA strands with specific structural properties. The direct analysis of genomic DNA and RNA in an array format without labeling or

Thiolated DNA-based chemistry and control in the structure and optical properties of plasmonic nanoparticles with ultrasmall interior nanogap.

PubMed

Oh, Jeong-Wook; Lim, Dong-Kwon; Kim, Gyeong-Hwan; Suh, Yung Doug; Nam, Jwa-Min

2014-10-08

The design, synthesis and control of plasmonic nanostructures, especially with ultrasmall plasmonically coupled nanogap (∼1 nm or smaller), are of significant interest and importance in chemistry, nanoscience, materials science, optics and nanobiotechnology. Here, we studied and established the thiolated DNA-based synthetic principles and methods in forming and controlling Au core-nanogap-Au shell structures [Au-nanobridged nanogap particles (Au-NNPs)] with various interior nanogap and Au shell structures. We found that differences in the binding affinities and modes among four different bases to Au core, DNA sequence, DNA grafting density and chemical reagents alter Au shell growth mechanism and interior nanogap-forming process on thiolated DNA-modified Au core. Importantly, poly A or poly C sequence creates a wider interior nanogap with a smoother Au shell, while poly T sequence results in a narrower interstitial interior gap with rougher Au shell, and on the basis of the electromagnetic field calculation and experimental results, we unraveled the relationships between the width of the interior plasmonic nanogap, Au shell structure, electromagnetic field and surface-enhanced Raman scattering. These principles and findings shown in this paper offer the fundamental basis for the thiolated DNA-based chemistry in forming and controlling metal nanostructures with ∼1 nm plasmonic gap and insight in the optical properties of the plasmonic NNPs, and these plasmonic nanogap structures are useful as strong and controllable optical signal-generating nanoprobes.

DNA-Based Single-Molecule Electronics: From Concept to Function.

PubMed

Wang, Kun

2018-01-17

Beyond being the repository of genetic information, DNA is playing an increasingly important role as a building block for molecular electronics. Its inherent structural and molecular recognition properties render it a leading candidate for molecular electronics applications. The structural stability, diversity and programmability of DNA provide overwhelming freedom for the design and fabrication of molecular-scale devices. In the past two decades DNA has therefore attracted inordinate amounts of attention in molecular electronics. This review gives a brief survey of recent experimental progress in DNA-based single-molecule electronics with special focus on single-molecule conductance and I-V characteristics of individual DNA molecules. Existing challenges and exciting future opportunities are also discussed.

DNA-Based Single-Molecule Electronics: From Concept to Function

PubMed Central

2018-01-01

Beyond being the repository of genetic information, DNA is playing an increasingly important role as a building block for molecular electronics. Its inherent structural and molecular recognition properties render it a leading candidate for molecular electronics applications. The structural stability, diversity and programmability of DNA provide overwhelming freedom for the design and fabrication of molecular-scale devices. In the past two decades DNA has therefore attracted inordinate amounts of attention in molecular electronics. This review gives a brief survey of recent experimental progress in DNA-based single-molecule electronics with special focus on single-molecule conductance and I–V characteristics of individual DNA molecules. Existing challenges and exciting future opportunities are also discussed. PMID:29342091

Minimalist Approach to Complexity: Templating the Assembly of DNA Tile Structures with Sequentially Grown Input Strands.

PubMed

Lau, Kai Lin; Sleiman, Hanadi F

2016-07-26

Given its highly predictable self-assembly properties, DNA has proven to be an excellent template toward the design of functional materials. Prominent examples include the remarkable complexity provided by DNA origami and single-stranded tile (SST) assemblies, which require hundreds of unique component strands. However, in many cases, the majority of the DNA assembly is purely structural, and only a small "working area" needs to be aperiodic. On the other hand, extended lattices formed by DNA tile motifs require only a few strands; but they suffer from lack of size control and limited periodic patterning. To overcome these limitations, we adopt a templation strategy, where an input strand of DNA dictates the size and patterning of resultant DNA tile structures. To prepare these templating input strands, a sequential growth technique developed in our lab is used, whereby extended DNA strands of defined sequence and length may be generated simply by controlling their order of addition. With these, we demonstrate the periodic patterning of size-controlled double-crossover (DX) and triple-crossover (TX) tile structures, as well as intentionally designed aperiodicity of a DX tile structure. As such, we are able to prepare size-controlled DNA structures featuring aperiodicity only where necessary with exceptional economy and efficiency.

Supramolecular Hydrogels Based on DNA Self-Assembly.

PubMed

Shao, Yu; Jia, Haoyang; Cao, Tianyang; Liu, Dongsheng

2017-04-18

Extracellular matrix (ECM) provides essential supports three dimensionally to the cells in living organs, including mechanical support and signal, nutrition, oxygen, and waste transportation. Thus, using hydrogels to mimic its function has attracted much attention in recent years, especially in tissue engineering, cell biology, and drug screening. However, a hydrogel system that can merit all parameters of the natural ECM is still a challenge. In the past decade, deoxyribonucleic acid (DNA) has arisen as an outstanding building material for the hydrogels, as it has unique properties compared to most synthetic or natural polymers, such as sequence designability, precise recognition, structural rigidity, and minimal toxicity. By simple attachment to polymers as a side chain, DNA has been widely used as cross-links in hydrogel preparation. The formed secondary structures could confer on the hydrogel designable responsiveness, such as response to temperature, pH, metal ions, proteins, DNA, RNA, and small signal molecules like ATP. Moreover, single or multiple DNA restriction enzyme sites could be incorporated into the hydrogels by sequence design and greatly expand the latitude of their responses. Compared with most supramolecular hydrogels, these DNA cross-linked hydrogels could be relatively strong and easily adjustable via sequence variation, but it is noteworthy that these hydrogels still have excellent thixotropic properties and could be easily injected through a needle. In addition, the quick formation of duplex has also enabled the multilayer three-dimensional injection printing of living cells with the hydrogel as matrix. When the matrix is built purely by DNA assembly structures, the hydrogel inherits all the previously described characteristics; however, the long persistence length of DNA structures excluded the small size meshes of the network and made the hydrogel permeable to nutrition for cell proliferation. This unique property greatly expands the cell viability in the three-dimensional matrix to several weeks and also provides an easy way to prepare interpenetrating double network materials. In this Account, we outline the stream of hydrogels based on DNA self-assembly and discuss the mechanism that brings outstanding properties to the materials. Unlike most reported hydrogel systems, the all-in-one character of the DNA hydrogel avoids the "cask effect" in the properties. We believe the hydrogel will greatly benefit cell behavior studies especially in the following aspects: (1) stem cell differentiation can be studied with solely tunable mechanical strength of the matrix; (2) the dynamic nature of the network can allow cell migration through the hydrogel, which will help to build a more realistic model to observe the migration of cancer cells in vivo; (3) combination with rapidly developing three-dimension printing technology, the hydrogel will boost the construction of three-dimensional tissues and artificial organs.

Adaptive resolution simulation of oligonucleotides

NASA Astrophysics Data System (ADS)

Netz, Paulo A.; Potestio, Raffaello; Kremer, Kurt

2016-12-01

Nucleic acids are characterized by a complex hierarchical structure and a variety of interaction mechanisms with other molecules. These features suggest the need of multiscale simulation methods in order to grasp the relevant physical properties of deoxyribonucleic acid (DNA) and RNA using in silico experiments. Here we report an implementation of a dual-resolution modeling of a DNA oligonucleotide in physiological conditions; in the presented setup only the nucleotide molecule and the solvent and ions in its proximity are described at the atomistic level; in contrast, the water molecules and ions far from the DNA are represented as computationally less expensive coarse-grained particles. Through the analysis of several structural and dynamical parameters, we show that this setup reliably reproduces the physical properties of the DNA molecule as observed in reference atomistic simulations. These results represent a first step towards a realistic multiscale modeling of nucleic acids and provide a quantitatively solid ground for their simulation using dual-resolution methods.

DNA origami compliant nanostructures with tunable mechanical properties.

PubMed

Zhou, Lifeng; Marras, Alexander E; Su, Hai-Jun; Castro, Carlos E

2014-01-28

DNA origami enables fabrication of precise nanostructures by programming the self-assembly of DNA. While this approach has been used to make a variety of complex 2D and 3D objects, the mechanical functionality of these structures is limited due to their rigid nature. We explore the fabrication of deformable, or compliant, objects to establish a framework for mechanically functional nanostructures. This compliant design approach is used in macroscopic engineering to make devices including sensors, actuators, and robots. We build compliant nanostructures by utilizing the entropic elasticity of single-stranded DNA (ssDNA) to locally bend bundles of double-stranded DNA into bent geometries whose curvature and mechanical properties can be tuned by controlling the length of ssDNA strands. We demonstrate an ability to achieve a wide range of geometries by adjusting a few strands in the nanostructure design. We further developed a mechanical model to predict both geometry and mechanical properties of our compliant nanostructures that agrees well with experiments. Our results provide a basis for the design of mechanically functional DNA origami devices and materials.

3D DNA Crystals and Nanotechnology

DOE Office of Scientific and Technical Information (OSTI.GOV)

Paukstelis, Paul; Seeman, Nadrian

DNA's molecular recognition properties have made it one of the most widely used biomacromolecular construction materials. The programmed assembly of DNA oligonucleotides has been used to create complex 2D and 3D self-assembled architectures and to guide the assembly of other molecules. The origins of DNA nanotechnology are rooted in the goal of assembling DNA molecules into designed periodic arrays, i.e., crystals. Here, we highlight several DNA crystal structures, the progress made in designing DNA crystals, and look at the current prospects and future directions of DNA crystals in nanotechnology.

3D DNA Crystals and Nanotechnology

DOE PAGES

Paukstelis, Paul; Seeman, Nadrian

2016-08-18

DNA's molecular recognition properties have made it one of the most widely used biomacromolecular construction materials. The programmed assembly of DNA oligonucleotides has been used to create complex 2D and 3D self-assembled architectures and to guide the assembly of other molecules. The origins of DNA nanotechnology are rooted in the goal of assembling DNA molecules into designed periodic arrays, i.e., crystals. Here, we highlight several DNA crystal structures, the progress made in designing DNA crystals, and look at the current prospects and future directions of DNA crystals in nanotechnology.

Three-dimensional structural dynamics and fluctuations of DNA-nanogold conjugates by individual-particle electron tomography

DOE PAGES

Zhang, Lei; Lei, Dongsheng; Smith, Jessica M.; ...

2016-03-30

DNA base pairing has been used for many years to direct the arrangement of inorganic nanocrystals into small groupings and arrays with tailored optical and electrical properties. The control of DNA-mediated assembly depends crucially on a better understanding of three-dimensional structure of DNA-nanocrystal-hybridized building blocks. Existing techniques do not allow for structural determination of these flexible and heterogeneous samples. Here we report cryo-electron microscopy and negative-staining electron tomography approaches to image, and three-dimensionally reconstruct a single DNA-nanogold conjugate, an 84-bp double-stranded DNA with two 5-nm nanogold particles for potential substrates in plasmon-coupling experiments. By individual-particle electron tomography reconstruction, we obtainmore » 14 density maps at ~ 2-nm resolution . Using these maps as constraints, we derive 14 conformations of dsDNA by molecular dynamics simulations. The conformational variation is consistent with that from liquid solution, suggesting that individual-particle electron tomography could be an expected approach to study DNA-assembling and flexible protein structure and dynamics.« less

Superstatistical model of bacterial DNA architecture

NASA Astrophysics Data System (ADS)

Bogachev, Mikhail I.; Markelov, Oleg A.; Kayumov, Airat R.; Bunde, Armin

2017-02-01

Understanding the physical principles that govern the complex DNA structural organization as well as its mechanical and thermodynamical properties is essential for the advancement in both life sciences and genetic engineering. Recently we have discovered that the complex DNA organization is explicitly reflected in the arrangement of nucleotides depicted by the universal power law tailed internucleotide interval distribution that is valid for complete genomes of various prokaryotic and eukaryotic organisms. Here we suggest a superstatistical model that represents a long DNA molecule by a series of consecutive ~150 bp DNA segments with the alternation of the local nucleotide composition between segments exhibiting long-range correlations. We show that the superstatistical model and the corresponding DNA generation algorithm explicitly reproduce the laws governing the empirical nucleotide arrangement properties of the DNA sequences for various global GC contents and optimal living temperatures. Finally, we discuss the relevance of our model in terms of the DNA mechanical properties. As an outlook, we focus on finding the DNA sequences that encode a given protein while simultaneously reproducing the nucleotide arrangement laws observed from empirical genomes, that may be of interest in the optimization of genetic engineering of long DNA molecules.

Layered graphene-mica substrates induce melting of DNA origami

NASA Astrophysics Data System (ADS)

Green, Nathaniel S.; Pham, Phi H. Q.; Crow, Daniel T.; Burke, Peter J.; Norton, Michael L.

2018-04-01

Monolayer graphene supported on mica substrates induce melting of cross-shaped DNA origami. This behavior can be contrasted with the case of origami on graphene on graphite, where an expansion or partially re-organized structure is observed. On mica, only well-formed structures are observed. Comparison of the morphological differences observed for these probes after adsorption on these substrates provides insights into the sensitivity of DNA based nanostructures to the properties of the graphene monolayer, as modified by its substrate.

Size-expanded DNA bases: An ab-initio study of their structural and electronic properties

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fuentes-Cabrera, Miguel A; Sumpter, Bobby G; Wells, Jack C

2005-01-01

The size-expanded DNA bases, xA, xC, xG, and xT, are benzo-homologue forms of the natural DNA bases; i.e., their structure can be seen as the fusion of a natural base and a benzene ring. Recently, a variety of DNAs, known as xDNAs, have been synthesized in which size-expanded and natural bases are paired. In this paper we use second-order Moeller-Plesset perturbation theory and density functional theory to investigate the structural and electronic properties of xA, xC, xG, and xT and their natural counterparts. We find that whereas natural and size-expanded bases have both nonplanar amino groups the latter have alsomore » nonplanar aromatic rings. When density functional theory is used to investigate the electronic properties of size-expanded and natural bases, it is found that the HOMO-LUMO gap of the size-expanded bases is smaller than that of the natural bases. Also, xG should be easier to oxidize than G.« less

Self-assembly of hierarchically ordered structures in DNA nanotube systems

NASA Astrophysics Data System (ADS)

Glaser, Martin; Schnauß, Jörg; Tschirner, Teresa; Schmidt, B. U. Sebastian; Moebius-Winkler, Maximilian; Käs, Josef A.; Smith, David M.

2016-05-01

The self-assembly of molecular and macromolecular building blocks into organized patterns is a complex process found in diverse systems over a wide range of size and time scales. The formation of star- or aster-like configurations, for example, is a common characteristic in solutions of polymers or other molecules containing multi-scaled, hierarchical assembly processes. This is a recurring phenomenon in numerous pattern-forming systems ranging from cellular constructs to solutions of ferromagnetic colloids or synthetic plastics. To date, however, it has not been possible to systematically parameterize structural properties of the constituent components in order to study their influence on assembled states. Here, we circumvent this limitation by using DNA nanotubes with programmable mechanical properties as our basic building blocks. A small set of DNA oligonucleotides can be chosen to hybridize into micron-length DNA nanotubes with a well-defined circumference and stiffness. The self-assembly of these nanotubes to hierarchically ordered structures is driven by depletion forces caused by the presence of polyethylene glycol. This trait allowed us to investigate self-assembly effects while maintaining a complete decoupling of density, self-association or bundling strength, and stiffness of the nanotubes. Our findings show diverse ranges of emerging structures including heterogeneous networks, aster-like structures, and densely bundled needle-like structures, which compare to configurations found in many other systems. These show a strong dependence not only on concentration and bundling strength, but also on the underlying mechanical properties of the nanotubes. Similar network architectures to those caused by depletion forces in the low-density regime are obtained when an alternative hybridization-based bundling mechanism is employed to induce self-assembly in an isotropic network of pre-formed DNA nanotubes. This emphasizes the universal effect inevitable attractive forces in crowded environments have on systems of self-assembling soft matter, which should be considered for macromolecular structures applied in crowded systems such as cells.

Structure–Function Studies of DNA Polymerase λ

PubMed Central

2015-01-01

DNA polymerase λ (pol λ) functions in DNA repair with its main roles considered to be filling short gaps during repair of double-strand breaks by nonhomologous end joining and during base excision repair. As indicated by structural and biochemical studies over the past 10 years, pol λ shares many common properties with other family X siblings (pol β, pol μ, and terminal deoxynucleotidyl transferase) but also has unique structural features that determine its specific functions. In this review, we consider how structural studies over the past decade furthered our understanding of the behavior and biological roles of pol λ. PMID:24716527

Comparing Charge Transport in Oligonucleotides: RNA:DNA Hybrids and DNA Duplexes.

PubMed

Li, Yuanhui; Artés, Juan M; Qi, Jianqing; Morelan, Ian A; Feldstein, Paul; Anantram, M P; Hihath, Joshua

2016-05-19

Understanding the electronic properties of oligonucleotide systems is important for applications in nanotechnology, biology, and sensing systems. Here the charge-transport properties of guanine-rich RNA:DNA hybrids are compared to double-stranded DNA (dsDNA) duplexes with identical sequences. The conductance of the RNA:DNA hybrids is ∼10 times higher than the equivalent dsDNA, and conformational differences are determined to be the primary reason for this difference. The conductance of the RNA:DNA hybrids is also found to decrease more rapidly than dsDNA when the length is increased. Ab initio electronic structure and Green's function-based density of states calculations demonstrate that these differences arise because the energy levels are more spatially distributed in the RNA:DNA hybrid but that the number of accessible hopping sites is smaller. These combination results indicate that a simple hopping model that treats each individual guanine as a hopping site is insufficient to explain both a higher conductance and β value for RNA:DNA hybrids, and larger delocalization lengths must be considered.

An introduction to the mechanics of DNA.

PubMed

Travers, A A; Thompson, J M T

2004-07-15

This article gives an overview of recent research on the mechanical properties and spatial deformations of the DNA molecule. Globally the molecule behaves like a uniform elastic rod, and its twisting and writhing govern its compaction and packaging within a cell. Meanwhile high mechanical stresses can induce structural transitions of DNA giving, for example, a phase diagram in the space of the applied tension and torque. Locally, the mechanical properties vary according to the local sequence organization. These variations play a vital role in the biological functioning of the molecule.

Low-temperature 1H NMR spectroscopic study of hydration properties of a hybrid system based on nanosilica, DNA and doxorubicin in the presence of C60 fullerene

NASA Astrophysics Data System (ADS)

Turov, V. V.; Prylutskyy, Yu. I.; Ugnivenko, A. P.; Barvinchenko, V. N.; Krupskaya, T. V.; Tsierkezos, N. G.; Ritter, U.

2014-03-01

The structure of hydrate cover layers of SiO2-DNA-Dox (where Dox: doxorubicin) and SiO2-DNA-Dox-C60 fullerene hybrids was studied by means of low-temperature 1H NMR spectroscopy in tetrachloromethane. The hydration properties of SiO2-DNA-Dox nanomaterials combined with fullerenes and their derivatives are extremely important for their further use as therapeutics in cancer treatment and for safety reasons. The findings reveal that the hydration properties of the hybrids differ from those of the solid DNA particulates or SiO2-DNA systems due to the existence of different types of water clusters, namely the weakly (WAW) and strongly associated water (SAW) clusters. For SAW clusters the radial distributions as well as the distributions of change in Gibbs free energy due to adsorptive interactions at the surfaces of the investigated systems were obtained.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Egli, Martin; Pallan, Pradeep S.; Pattanayek, Rekha

An experimental rationalization of the structure type encountered in DNA and RNA by systematically investigating the chemical and physical properties of alternative nucleic acids has identified systems with a variety of sugar-phosphate backbones that are capable of Watson-Crick base pairing and in some cases cross-pairing with the natural nucleic acids. The earliest among the model systems tested to date, (4{prime} {yields} 6{prime})-linked oligo(2{prime},3{prime}-dideoxy-{beta}-d-glucopyranosyl)nucleotides or homo-DNA, shows stable self-pairing, but the pairing rules for the four natural bases are not the same as those in DNA. However, a complete interpretation and understanding of the properties of the hexapyranosyl (4{prime} {yields} 6{prime})more » family of nucleic acids has been impeded until now by the lack of detailed 3D-structural data. We have determined the crystal structure of a homo-DNA octamer. It reveals a weakly twisted right-handed duplex with a strong inclination between the hexose-phosphate backbones and base-pair axes, and highly irregular values for helical rise and twist at individual base steps. The structure allows a rationalization of the inability of allo-, altro-, and glucopyranosyl-based oligonucleotides to form stable pairing systems.« less

The Dynamic Interplay Between DNA Topoisomerases and DNA Topology.

PubMed

Seol, Yeonee; Neuman, Keir C

2016-09-01

Topological properties of DNA influence its structure and biochemical interactions. Within the cell DNA topology is constantly in flux. Transcription and other essential processes including DNA replication and repair, alter the topology of the genome, while introducing additional complications associated with DNA knotting and catenation. These topological perturbations are counteracted by the action of topoisomerases, a specialized class of highly conserved and essential enzymes that actively regulate the topological state of the genome. This dynamic interplay among DNA topology, DNA processing enzymes, and DNA topoisomerases, is a pervasive factor that influences DNA metabolism in vivo . Building on the extensive structural and biochemical characterization over the past four decades that established the fundamental mechanistic basis of topoisomerase activity, the unique roles played by DNA topology in modulating and influencing the activity of topoisomerases have begun to be explored. In this review we survey established and emerging DNA topology dependent protein-DNA interactions with a focus on in vitro measurements of the dynamic interplay between DNA topology and topoisomerase activity.

The dynamic interplay between DNA topoisomerases and DNA topology.

PubMed

Seol, Yeonee; Neuman, Keir C

2016-11-01

Topological properties of DNA influence its structure and biochemical interactions. Within the cell, DNA topology is constantly in flux. Transcription and other essential processes, including DNA replication and repair, not only alter the topology of the genome but also introduce additional complications associated with DNA knotting and catenation. These topological perturbations are counteracted by the action of topoisomerases, a specialized class of highly conserved and essential enzymes that actively regulate the topological state of the genome. This dynamic interplay among DNA topology, DNA processing enzymes, and DNA topoisomerases is a pervasive factor that influences DNA metabolism in vivo. Building on the extensive structural and biochemical characterization over the past four decades that has established the fundamental mechanistic basis of topoisomerase activity, scientists have begun to explore the unique roles played by DNA topology in modulating and influencing the activity of topoisomerases. In this review we survey established and emerging DNA topology-dependent protein-DNA interactions with a focus on in vitro measurements of the dynamic interplay between DNA topology and topoisomerase activity.

Stimuli-Responsive DNA-Based Hydrogels: From Basic Principles to Applications.

PubMed

Kahn, Jason S; Hu, Yuwei; Willner, Itamar

2017-04-18

The base sequence of nucleic acids encodes structural and functional information into the DNA biopolymer. External stimuli such as metal ions, pH, light, or added nucleic acid fuel strands provide triggers to reversibly switch nucleic acid structures such as metal-ion-bridged duplexes, i-motifs, triplex nucleic acids, G-quadruplexes, or programmed double-stranded hybrids of oligonucleotides (DNA). The signal-triggered oligonucleotide structures have been broadly applied to develop switchable DNA nanostructures and DNA machines, and these stimuli-responsive assemblies provide functional scaffolds for the rapidly developing area of DNA nanotechnology. Stimuli-responsive hydrogels undergoing signal-triggered hydrogel-to-solution transitions or signal-controlled stiffness changes attract substantial interest as functional matrices for controlled drug delivery, materials exhibiting switchable mechanical properties, acting as valves or actuators, and "smart" materials for sensing and information processing. The integration of stimuli-responsive oligonucleotides with hydrogel-forming polymers provides versatile means to exploit the functional information encoded in the nucleic acid sequences to yield stimuli-responsive hydrogels exhibiting switchable physical, structural, and chemical properties. Stimuli-responsive DNA-based nucleic acid structures are integrated in acrylamide polymer chains and reversible, switchable hydrogel-to-solution transitions of the systems are demonstrated by applying external triggers, such as metal ions, pH-responsive strands, G-quadruplex, and appropriate counter triggers that bridge and dissociate the polymer chains. By combining stimuli-responsive nucleic acid bridges with thermosensitive poly(N-isopropylacrylamide) (pNIPAM) chains, systems undergoing reversible solution ↔ hydrogel ↔ solid transitions are demonstrated. Specifically, by bridging acrylamide polymer chains by two nucleic acid functionalities, where one type of bridging unit provides a stimuli-responsive element and the second unit acts as internal "bridging memory", shape-memory hydrogels undergoing reversible and switchable transitions between shaped hydrogels and shapeless quasi-liquid states are demonstrated. By using stimuli-responsive hydrogel cross-linking units that can assemble the bridging units by two different input signals, the orthogonally-triggered functions of the shape-memory were shown. Furthermore, a versatile approach to assemble stimuli-responsive DNA-based acrylamide hydrogel films on surfaces is presented. The method involves the activation of the hybridization chain-reaction (HCR) by a surface-confined promoter strand, in the presence of acrylamide chains modified with two DNA hairpin structures and appropriate stimuli-responsive tethers. The resulting hydrogel-modified surfaces revealed switchable stiffness properties and signal-triggered catalytic functions. By applying the method to assemble the hydrogel microparticles, substrate-loaded, stimuli-responsive microcapsules are prepared. The signal-triggered DNA-based hydrogel microcapsules are applied as drug carriers for controlled release. The different potential applications and future perspectives of stimuli responsive hydrogels are discussed. Specifically, the use of these smart materials and assemblies as carriers for controlled drug release and as shape-memory matrices for information storage and inscription and the use of surface-confined stimuli-responsive hydrogels, exhibiting switchable stiffness properties, for catalysis and controlled growth of cells are discussed.

Solution structure and DNA-binding properties of the C-terminal domain of UvrC from E.coli

PubMed Central

Singh, S.; Folkers, G.E.; Bonvin, A.M.J.J.; Boelens, R.; Wechselberger, R.; Niztayev, A.; Kaptein, R.

2002-01-01

The C-terminal domain of the UvrC protein (UvrC CTD) is essential for 5′ incision in the prokaryotic nucleotide excision repair process. We have determined the three-dimensional structure of the UvrC CTD using heteronuclear NMR techniques. The structure shows two helix–hairpin–helix (HhH) motifs connected by a small connector helix. The UvrC CTD is shown to mediate structure-specific DNA binding. The domain binds to a single-stranded–double-stranded junction DNA, with a strong specificity towards looped duplex DNA that contains at least six unpaired bases per loop (‘bubble DNA’). Using chemical shift perturbation experiments, the DNA-binding surface is mapped to the first hairpin region encompassing the conserved glycine–valine–glycine residues followed by lysine–arginine–arginine, a positively charged surface patch and the second hairpin region consisting of glycine–isoleucine–serine. A model for the protein– DNA complex is proposed that accounts for this specificity. PMID:12426397

Chemical biology on the genome.

PubMed

Balasubramanian, Shankar

2014-08-15

In this article I discuss studies towards understanding the structure and function of DNA in the context of genomes from the perspective of a chemist. The first area I describe concerns the studies that led to the invention and subsequent development of a method for sequencing DNA on a genome scale at high speed and low cost, now known as Solexa/Illumina sequencing. The second theme will feature the four-stranded DNA structure known as a G-quadruplex with a focus on its fundamental properties, its presence in cellular genomic DNA and the prospects for targeting such a structure in cels with small molecules. The final topic for discussion is naturally occurring chemically modified DNA bases with an emphasis on chemistry for decoding (or sequencing) such modifications in genomic DNA. The genome is a fruitful topic to be further elucidated by the creation and application of chemical approaches. Copyright © 2014 Elsevier Ltd. All rights reserved.

An experimentally-informed coarse-grained 3-site-per-nucleotide model of DNA: Structure, thermodynamics, and dynamics of hybridization

PubMed Central

Hinckley, Daniel M.; Freeman, Gordon S.; Whitmer, Jonathan K.; de Pablo, Juan J.

2013-01-01

A new 3-Site-Per-Nucleotide coarse-grained model for DNA is presented. The model includes anisotropic potentials between bases involved in base stacking and base pair interactions that enable the description of relevant structural properties, including the major and minor grooves. In an improvement over available coarse-grained models, the correct persistence length is recovered for both ssDNA and dsDNA, allowing for simulation of non-canonical structures such as hairpins. DNA melting temperatures, measured for duplexes and hairpins by integrating over free energy surfaces generated using metadynamics simulations, are shown to be in quantitative agreement with experiment for a variety of sequences and conditions. Hybridization rate constants, calculated using forward-flux sampling, are also shown to be in good agreement with experiment. The coarse-grained model presented here is suitable for use in biological and engineering applications, including nucleosome positioning and DNA-templated engineering. PMID:24116642

T7 RNA polymerase non-specifically transcribes and induces disassembly of DNA nanostructures

PubMed Central

Schaffter, Samuel W; Green, Leopold N; Schneider, Joanna; Subramanian, Hari K K; Schulman, Rebecca

2018-01-01

Abstract The use of proteins that bind and catalyze reactions with DNA alongside DNA nanostructures has broadened the functionality of DNA devices. DNA binding proteins have been used to specifically pattern and tune structural properties of DNA nanostructures and polymerases have been employed to directly and indirectly drive structural changes in DNA structures and devices. Despite these advances, undesired and poorly understood interactions between DNA nanostructures and proteins that bind DNA continue to negatively affect the performance and stability of DNA devices used in conjunction with enzymes. A better understanding of these undesired interactions will enable the construction of robust DNA nanostructure-enzyme hybrid systems. Here, we investigate the undesired disassembly of DNA nanotubes in the presence of viral RNA polymerases (RNAPs) under conditions used for in vitro transcription. We show that nanotubes and individual nanotube monomers (tiles) are non-specifically transcribed by T7 RNAP, and that RNA transcripts produced during non-specific transcription disassemble the nanotubes. Disassembly requires a single-stranded overhang on the nanotube tiles where transcripts can bind and initiate disassembly through strand displacement, suggesting that single-stranded domains on other DNA nanostructures could cause unexpected interactions in the presence of viral RNA polymerases. PMID:29718412

T7 RNA polymerase non-specifically transcribes and induces disassembly of DNA nanostructures.

PubMed

Schaffter, Samuel W; Green, Leopold N; Schneider, Joanna; Subramanian, Hari K K; Schulman, Rebecca; Franco, Elisa

2018-06-01

The use of proteins that bind and catalyze reactions with DNA alongside DNA nanostructures has broadened the functionality of DNA devices. DNA binding proteins have been used to specifically pattern and tune structural properties of DNA nanostructures and polymerases have been employed to directly and indirectly drive structural changes in DNA structures and devices. Despite these advances, undesired and poorly understood interactions between DNA nanostructures and proteins that bind DNA continue to negatively affect the performance and stability of DNA devices used in conjunction with enzymes. A better understanding of these undesired interactions will enable the construction of robust DNA nanostructure-enzyme hybrid systems. Here, we investigate the undesired disassembly of DNA nanotubes in the presence of viral RNA polymerases (RNAPs) under conditions used for in vitro transcription. We show that nanotubes and individual nanotube monomers (tiles) are non-specifically transcribed by T7 RNAP, and that RNA transcripts produced during non-specific transcription disassemble the nanotubes. Disassembly requires a single-stranded overhang on the nanotube tiles where transcripts can bind and initiate disassembly through strand displacement, suggesting that single-stranded domains on other DNA nanostructures could cause unexpected interactions in the presence of viral RNA polymerases.

Plasmonic nanostructures through DNA-assisted lithography

PubMed Central

Shen, Boxuan; Linko, Veikko; Tapio, Kosti; Pikker, Siim; Lemma, Tibebe; Gopinath, Ashwin; Gothelf, Kurt V.; Kostiainen, Mauri A.; Toppari, J. Jussi

2018-01-01

Programmable self-assembly of nucleic acids enables the fabrication of custom, precise objects with nanoscale dimensions. These structures can be further harnessed as templates to build novel materials such as metallic nanostructures, which are widely used and explored because of their unique optical properties and their potency to serve as components of novel metamaterials. However, approaches to transfer the spatial information of DNA constructions to metal nanostructures remain a challenge. We report a DNA-assisted lithography (DALI) method that combines the structural versatility of DNA origami with conventional lithography techniques to create discrete, well-defined, and entirely metallic nanostructures with designed plasmonic properties. DALI is a parallel, high-throughput fabrication method compatible with transparent substrates, thus providing an additional advantage for optical measurements, and yields structures with a feature size of ~10 nm. We demonstrate its feasibility by producing metal nanostructures with a chiral plasmonic response and bowtie-shaped nanoantennas for surface-enhanced Raman spectroscopy. We envisage that DALI can be generalized to large substrates, which would subsequently enable scale-up production of diverse metallic nanostructures with tailored plasmonic features. PMID:29423446

Animal Mitochondrial DNA Replication

PubMed Central

Ciesielski, Grzegorz L.; Oliveira, Marcos T.; Kaguni, Laurie S.

2016-01-01

Recent advances in the field of mitochondrial DNA (mtDNA) replication highlight the diversity of both the mechanisms utilized and the structural and functional organization of the proteins at mtDNA replication fork, despite the simplicity of the animal mtDNA genome. DNA polymerase γ, mtDNA helicase and mitochondrial single-stranded DNA-binding protein- the key replisome proteins, have evolved distinct structural features and biochemical properties. These appear to be correlated with mtDNA genomic features in different metazoan taxa and with their modes of DNA replication, although a substantial integrative research is warranted to establish firmly these links. To date, several modes of mtDNA replication have been described for animals: rolling circle, theta, strand-displacement, and RITOLS/bootlace. Resolution of a continuing controversy relevant to mtDNA replication in mammals/vertebrates will have a direct impact on the mechanistic interpretation of mtDNA-related human diseases. Here we review these subjects, integrating earlier and recent data to provide a perspective on the major challenges for future research. PMID:27241933

Size-Expanded yDNA bases: An Ab Initio Study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fuentes-Cabrera, Miguel A; Sumpter, Bobby G; Lipkowski, Pawel

2006-01-01

xDNA and yDNA are new classes of synthetic nucleic acids characterized by having base-pairs with one of the bases larger than the natural congeners. Here these larger bases are called x- and y-bases. We recently investigated and reported the structural and electronic properties of the x-bases (Fuentes-Cabrera et al. J. Phys. Chem. B 2005, 109, 21135-21139). Here we extend this study by investigating the structure and electronic properties of the y-bases. These studies are framed within our interest that xDNA and yDNA could function as nanowires, for they could have smaller HOMO-LUMO gaps than natural DNA. The limited amount ofmore » experimental structural data in these synthetic duplexes makes it necessary to first understand smaller models and, subsequently, to use that information to build larger models. In this paper, we report the results on the chemical and electronic structure of the y-bases. In particular, we predict that the y-bases have smaller HOMO-LUMO gaps than their natural congeners, which is an encouraging result for it indicates that yDNA could have a smaller HOMO-LUMO gap than natural DNA. Also, we predict that the y-bases are less planar than the natural ones. Particularly interesting are our results corresponding to yG. Our studies show that yG is unstable because it is less aromatic and has a Coulombic repulsion that involves the amino group, as compared with a more stable tautomer. However, yG has a very small HOMO-LUMO gap, the smallest of all the size-expanded bases we have considered. The results of this study provide useful information that may allow the synthesis of an yG-mimic that is stable and has a small HOMO-LUMO gap.« less

Crystal Structure of the GRAS Domain of SCARECROW-LIKE7 in Oryza sativa

PubMed Central

Li, Shengping; Zhao, Yanhe; Zhao, Zheng; Wu, Xiuling; Sun, Lifang; Liu, Qingsong; Wu, Yunkun

2016-01-01

GRAS proteins belong to a plant-specific protein family with many members and play essential roles in plant growth and development, functioning primarily in transcriptional regulation. Proteins in the family are minimally defined as containing the conserved GRAS domain. Here, we determined the structure of the GRAS domain of Os-SCL7 from rice (Oryza sativa) to 1.82 Å. The structure includes cap and core subdomains and elucidates the features of the conserved GRAS LRI, VHIID, LRII, PFYRE, and SAW motifs. The structure is a dimer, with a clear groove to accommodate double-stranded DNA. Docking a DNA segment into the groove to generate an Os-SCL7/DNA complex provides insight into the DNA binding mechanism of GRAS proteins. Furthermore, the in vitro DNA binding property of Os-SCL7 and model-defined recognition residues are assessed by electrophoretic mobility shift analysis and mutagenesis assays. These studies reveal the structure and preliminary DNA interaction mechanisms of GRAS proteins and open the door to in-depth investigation and understanding of the individual pathways in which they play important roles. PMID:27081181

The structure and DNA-binding properties of Mgm101 from a yeast with a linear mitochondrial genome.

PubMed

Pevala, Vladimír; Truban, Dominika; Bauer, Jacob A; Košťan, Július; Kunová, Nina; Bellová, Jana; Brandstetter, Marlene; Marini, Victoria; Krejčí, Lumír; Tomáška, Ľubomír; Nosek, Jozef; Kutejová, Eva

2016-03-18

To study the mechanisms involved in the maintenance of a linear mitochondrial genome we investigated the biochemical properties of the recombination protein Mgm101 from Candida parapsilosis. We show that CpMgm101 complements defects associated with the Saccharomyces cerevisiae mgm101-1(ts) mutation and that it is present in both the nucleus and mitochondrial nucleoids of C. parapsilosis. Unlike its S. cerevisiae counterpart, CpMgm101 is associated with the entire nucleoid population and is able to bind to a broad range of DNA substrates in a non-sequence specific manner. CpMgm101 is also able to catalyze strand annealing and D-loop formation. CpMgm101 forms a roughly C-shaped trimer in solution according to SAXS. Electron microscopy of a complex of CpMgm101 with a model mitochondrial telomere revealed homogeneous, ring-shaped structures at the telomeric single-stranded overhangs. The DNA-binding properties of CpMgm101, together with its DNA recombination properties, suggest that it can play a number of possible roles in the replication of the mitochondrial genome and the maintenance of its telomeres. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Multivalent Lipid--DNA Complexes: Distinct DNA Compaction Regimes

NASA Astrophysics Data System (ADS)

Evans, Heather M.; Ahmad, A.; Ewert, K.; Safinya, C. R.

2004-03-01

Cationic liposomes (CL), while intrinsically advantageous in comparison to viruses, still have limited success for gene therapy and require more study. CL spontaneously self-assemble with DNA via counterion release, forming small particles approximately 200nm in diameter. X-ray diffraction reveals CL-DNA structures that are typically a multilamellar organization of lipids with DNA intercalated between the layers. We explore the structural properties of CL-DNA complexes formed with new multivalent lipids (Ewert et al, J. Med. Chem. 2002; 45:5023) that range from 2+ to 16+. Contrary to a simple prediction for the DNA interaxial spacing d_DNA based on a geometrical space-filling model, these lipids show dramatic DNA compaction, down to d_DNA ˜ 25 ÅVariations in the membrane charge density, σ _M, lead to distinct spacing regimes. We propose that this DNA condensation is controlled by a unique locking mechanism between the DNA double helix and the large, multivalent lipid head groups. Funded by NSF DMR-0203755 and NIH GM-59288.

Probing Human Telomeric DNA and RNA Topology and Ligand Binding in a Cellular Model by Using Responsive Fluorescent Nucleoside Probes.

PubMed

Manna, Sudeshna; Panse, Cornelia H; Sontakke, Vyankat A; Sangamesh, Sarangamath; Srivatsan, Seergazhi G

2017-08-17

The development of biophysical systems that enable an understanding of the structure and ligand-binding properties of G-quadruplex (GQ)-forming nucleic acid sequences in cells or models that mimic the cellular environment would be highly beneficial in advancing GQ-directed therapeutic strategies. Herein, the establishment of a biophysical platform to investigate the structure and recognition properties of human telomeric (H-Telo) DNA and RNA repeats in a cell-like confined environment by using conformation-sensitive fluorescent nucleoside probes and a widely used cellular model, bis(2-ethylhexyl) sodium sulfosuccinate reverse micelles (RMs), is described. The 2'-deoxy and ribonucleoside probes, composed of a 5-benzofuran uracil base analogue, faithfully report the aqueous micellar core through changes in their fluorescence properties. The nucleoside probes incorporated into different loops of H-Telo DNA and RNA oligonucleotide repeats are minimally perturbing and photophysically signal the formation of respective GQ structures in both aqueous buffer and RMs. Furthermore, these sensors enable a direct comparison of the binding affinity of a ligand to H-Telo DNA and RNA GQ structures in the bulk and confined environment of RMs. These results demonstrate that this combination of a GQ nucleoside probe and easy-to-handle RMs could provide new opportunities to study and devise screening-compatible assays in a cell-like environment to discover GQ binders of clinical potential. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

TOPICAL REVIEW: The physics of chromatin

NASA Astrophysics Data System (ADS)

Schiessel, Helmut

2003-05-01

Recent progress has been made in the understanding of the physical properties of chromatin - the dense complex of DNA and histone proteins that occupies the nuclei of plant and animal cells. Here I will focus on the two lowest levels of the hierarchy of DNA folding into the chromatin complex. (i) The nucleosome, the chromatin repeating unit consisting of a globular aggregate of eight histone proteins with the DNA wrapped around it: its overcharging, the DNA unwrapping transition, the 'sliding' of the octamer along the DNA. (ii) The 30 nm chromatin fibre, the necklace-like structure of nucleosomes connected via linker DNA: its geometry, its mechanical properties under stretching and its response to changing ionic conditions. I will stress that chromatin combines two seemingly contradictory features: (1) high compaction of DNA within the nuclear envelope and, at the same time, (2) accessibility to genes, promoter regions and gene regulatory sequences.

Label-free DNA biosensor based on resistance change of platinum nanoparticles assemblies.

PubMed

Skotadis, Evangelos; Voutyras, Konstantinos; Chatzipetrou, Marianneza; Tsekenis, Georgios; Patsiouras, Lampros; Madianos, Leonidas; Chatzandroulis, Stavros; Zergioti, Ioanna; Tsoukalas, Dimitris

2016-07-15

A novel nanoparticle based biosensor for the fast and simple detection of DNA hybridization events is presented. The sensor utilizes hybridized DNA's charge transport properties, combining them with metallic nanoparticle networks that act as nano-gapped electrodes. The DNA hybridization events can be detected by a significant reduction in the sensor's resistance due to the conductive bridging offered by hybridized DNA. By modifying the nanoparticle surface coverage, which can be controlled experimentally being a function of deposition time, and the structural properties of the electrodes, an optimized biosensor for the in situ detection of DNA hybridization events is ultimately fabricated. The fabricated biosensor exhibits a wide response range, covering four orders of magnitude, a limit of detection of 1nM and can detect a single base pair mismatch between probe and complementary DNA. Copyright © 2016 Elsevier B.V. All rights reserved.

Deformation of DNA molecules by hydrodynamic focusing

NASA Astrophysics Data System (ADS)

Wong, Pak Kin; Lee, Yi-Kuen; Ho, Chih-Ming

2003-12-01

The motion of a DNA molecule in a solvent flow reflects the deformation of a nano/microscale flexible mass spring structure by the forces exerted by the fluid molecules. The dynamics of individual molecules can reveal both fundamental properties of the DNA and basic understanding of the complex rheological properties of long-chain molecules. In this study, we report the dynamics of isolated DNA molecules under homogeneous extensional flow. Hydrodynamic focusing generates homogeneous extensional flow with uniform velocity in the transverse direction. The deformation of individual DNA molecules in the flow was visualized with video fluorescence microscopy. A coil stretch transition was observed when the Deborah number (De) is larger than 0.8. With a sudden stopping of the flow, the DNA molecule relaxes and recoils. The longest relaxation time of T2 DNA was determined to be 0.63 s when scaling viscosity to 0.9 cP.

Biochemical Regulatory Features of Activation-Induced Cytidine Deaminase Remain Conserved from Lampreys to Humans

PubMed Central

King, Justin J.; Amemiya, Chris T.; Hsu, Ellen

2017-01-01

ABSTRACT Activation-induced cytidine deaminase (AID) is a genome-mutating enzyme that initiates class switch recombination and somatic hypermutation of antibodies in jawed vertebrates. We previously described the biochemical properties of human AID and found that it is an unusual enzyme in that it exhibits binding affinities for its substrate DNA and catalytic rates several orders of magnitude higher and lower, respectively, than a typical enzyme. Recently, we solved the functional structure of AID and demonstrated that these properties are due to nonspecific DNA binding on its surface, along with a catalytic pocket that predominantly assumes a closed conformation. Here we investigated the biochemical properties of AID from a sea lamprey, nurse shark, tetraodon, and coelacanth: representative species chosen because their lineages diverged at the earliest critical junctures in evolution of adaptive immunity. We found that these earliest-diverged AID orthologs are active cytidine deaminases that exhibit unique substrate specificities and thermosensitivities. Significant amino acid sequence divergence among these AID orthologs is predicted to manifest as notable structural differences. However, despite major differences in sequence specificities, thermosensitivities, and structural features, all orthologs share the unusually high DNA binding affinities and low catalytic rates. This absolute conservation is evidence for biological significance of these unique biochemical properties. PMID:28716949

Solution properties of the archaeal CRISPR DNA repeat-binding homeodomain protein Cbp2

PubMed Central

Kenchappa, Chandra S.; Heidarsson, Pétur O.; Kragelund, Birthe B.; Garrett, Roger A.; Poulsen, Flemming M.

2013-01-01

Clustered regularly interspaced short palindromic repeats (CRISPR) form the basis of diverse adaptive immune systems directed primarily against invading genetic elements of archaea and bacteria. Cbp1 of the crenarchaeal thermoacidophilic order Sulfolobales, carrying three imperfect repeats, binds specifically to CRISPR DNA repeats and has been implicated in facilitating production of long transcripts from CRISPR loci. Here, a second related class of CRISPR DNA repeat-binding protein, denoted Cbp2, is characterized that contains two imperfect repeats and is found amongst members of the crenarchaeal thermoneutrophilic order Desulfurococcales. DNA repeat-binding properties of the Hyperthermus butylicus protein Cbp2Hb were characterized and its three-dimensional structure was determined by NMR spectroscopy. The two repeats generate helix-turn-helix structures separated by a basic linker that is implicated in facilitating high affinity DNA binding of Cbp2 by tethering the two domains. Structural studies on mutant proteins provide support for Cys7 and Cys28 enhancing high thermal stability of Cbp2Hb through disulphide bridge formation. Consistent with their proposed CRISPR transcriptional regulatory role, Cbp2Hb and, by inference, other Cbp1 and Cbp2 proteins are closely related in structure to homeodomain proteins with linked helix-turn-helix (HTH) domains, in particular the paired domain Pax and Myb family proteins that are involved in eukaryal transcriptional regulation. PMID:23325851

A domain in human EXOG converts apoptotic endonuclease to DNA-repair exonuclease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Szymanski, Michal R.; Yu, Wangsheng; Gmyrek, Aleksandra M.

Human EXOG (hEXOG) is a 5'-exonuclease that is crucial for mitochondrial DNA repair; the enzyme belongs to a nonspecific nuclease family that includes the apoptotic endonuclease EndoG. Here we report biochemical and structural studies of hEXOG, including structures in its apo form and in a complex with DNA at 1.81 and 1.85 Å resolution, respectively. A Wing domain, absent in other ββα-Me members, suppresses endonuclease activity, but confers on hEXOG a strong 5'-dsDNA exonuclease activity that precisely excises a dinucleotide using an intrinsic ‘tape-measure’. The symmetrical apo hEXOG homodimer becomes asymmetrical upon binding to DNA, providing a structural basis formore » how substrate DNA bound to one active site allosterically regulates the activity of the other. These properties of hEXOG suggest a pathway for mitochondrial BER that provides an optimal substrate for subsequent gap-filling synthesis by DNA polymerase γ.« less

Effects of sequence on DNA wrapping around histones

NASA Astrophysics Data System (ADS)

Ortiz, Vanessa

2011-03-01

A central question in biophysics is whether the sequence of a DNA strand affects its mechanical properties. In epigenetics, these are thought to influence nucleosome positioning and gene expression. Theoretical and experimental attempts to answer this question have been hindered by an inability to directly resolve DNA structure and dynamics at the base-pair level. In our previous studies we used a detailed model of DNA to measure the effects of sequence on the stability of naked DNA under bending. Sequence was shown to influence DNA's ability to form kinks, which arise when certain motifs slide past others to form non-native contacts. Here, we have now included histone-DNA interactions to see if the results obtained for naked DNA are transferable to the problem of nucleosome positioning. Different DNA sequences interacting with the histone protein complex are studied, and their equilibrium and mechanical properties are compared among themselves and with the naked case. NLM training grant to the Computation and Informatics in Biology and Medicine Training Program (NLM T15LM007359).

Less Pollen-Mediated Gene Flow for More Signatures of Glacial Lineages: Congruent Evidence from Balsam Fir cpDNA and mtDNA for Multiple Refugia in Eastern and Central North America

PubMed Central

Cinget, Benjamin; Gérardi, Sébastien; Beaulieu, Jean; Bousquet, Jean

2015-01-01

The phylogeographic structure and postglacial history of balsam fir (Abies balsamea), a transcontinental North American boreal conifer, was inferred using mitochondrial DNA (mtDNA) and chloroplast DNA (cpDNA) markers. Genetic structure among 107 populations (mtDNA data) and 75 populations (cpDNA data) was analyzed using Bayesian and genetic distance approaches. Population differentiation was high for mtDNA (dispersed by seeds only), but also for cpDNA (dispersed by seeds and pollen), indicating that pollen gene flow is more restricted in balsam fir than in other boreal conifers. Low cpDNA gene flow in balsam fir may relate to low pollen production due to the inherent biology of the species and populations being decimated by recurrent spruce budworm epidemics, and/or to low dispersal of pollen grains due to their peculiar structural properties. Accordingly, a phylogeographic structure was detected using both mtDNA and cpDNA markers and population structure analyses supported the existence of at least five genetically distinct glacial lineages in central and eastern North America. Four of these would originate from glacial refugia located south of the Laurentide ice sheet, while the last one would have persisted in the northern Labrador region. As expected due to reduced pollen-mediated gene flow, congruence between the geographic distribution of mtDNA and cpDNA lineages was higher than in other North American conifers. However, concordance was not complete, reflecting that restricted but nonetheless detectable cpDNA gene flow among glacial lineages occurred during the Holocene. As a result, new cpDNA and mtDNA genome combinations indicative of cytoplasmic genome capture were observed. PMID:25849816

Stability of DNA Origami Nanoarrays in Cell Lysate

PubMed Central

Mei, Qian; Wei, Xixi; Su, Fengyu; Liu, Yan; Youngbull, Cody; Johnson, Roger; Lindsay, Stuart; Yan, Hao; Meldrum, Deirdre

2012-01-01

Scaffolded DNA origami, a method to create self-assembled nanostructures with spatially addressable features, has recently been used to develop water-soluble molecular chips for label-free RNA detection, platforms for deterministic protein positioning, and single molecule reaction observatories. These applications highlight the possibility of exploiting the unique properties and biocompatibility of DNA nanostructures in live, cellular systems. Herein, we assembled several DNA origami nanostructures of differing shape, size and probes, and investigated their interaction with lysate obtained from various normal and cancerous cell lines. We separated and analyzed the origami–lysate mixtures using agarose gel electrophoresis and recovered the DNA structures for functional assay and subsequent microscopic examination. Our results demonstrate that DNA origami nanostructures are stable in cell lysate and can be easily separated from lysate mixtures, in contrast to natural, single- and double-stranded DNA. Atomic force microscope (AFM) and transmission electron microscope (TEM) images show that the DNA origami structures are fully intact after separation from cell lysates and hybridize to their targets, verifying the superior structural integrity and functionality of self-assembled DNA origami nanostructures relative to conventional oligonucleotides. The stability and functionality of DNA origami structures in cell lysate validate their use for biological applications, for example, as programmable molecular rafts or disease detection platforms. PMID:21366226

Exploring the Interactions of the Dietary Plant Flavonoids Fisetin and Naringenin with G-Quadruplex and Duplex DNA, Showing Contrasting Binding Behavior: Spectroscopic and Molecular Modeling Approaches.

PubMed

Bhattacharjee, Snehasish; Chakraborty, Sandipan; Sengupta, Pradeep K; Bhowmik, Sudipta

2016-09-01

Guanine-rich sequences have the propensity to fold into a four-stranded DNA structure known as a G-quadruplex (G4). G4 forming sequences are abundant in the promoter region of several oncogenes and become a key target for anticancer drug binding. Here we have studied the interactions of two structurally similar dietary plant flavonoids fisetin and naringenin with G4 as well as double stranded (duplex) DNA by using different spectroscopic and modeling techniques. Our study demonstrates the differential binding ability of the two flavonoids with G4 and duplex DNA. Fisetin more strongly interacts with parallel G4 structure than duplex DNA, whereas naringenin shows stronger binding affinity to duplex rather than G4 DNA. Molecular docking results also corroborate our spectroscopic results, and it was found that both of the ligands are stacked externally in the G4 DNA structure. C-ring planarity of the flavonoid structure appears to be a crucial factor for preferential G4 DNA recognition of flavonoids. The goal of this study is to explore the critical effects of small differences in the structure of closely similar chemical classes of such small molecules (flavonoids) which lead to the contrasting binding properties with the two different forms of DNA. The resulting insights may be expected to facilitate the designing of the highly selective G4 DNA binders based on flavonoid scaffolds.

The T4 Phage DNA Mimic Protein Arn Inhibits the DNA Binding Activity of the Bacterial Histone-like Protein H-NS*

PubMed Central

Ho, Chun-Han; Wang, Hao-Ching; Ko, Tzu-Ping; Chang, Yuan-Chih; Wang, Andrew H.-J.

2014-01-01

The T4 phage protein Arn (Anti restriction nuclease) was identified as an inhibitor of the restriction enzyme McrBC. However, until now its molecular mechanism remained unclear. In the present study we used structural approaches to investigate biological properties of Arn. A structural analysis of Arn revealed that its shape and negative charge distribution are similar to dsDNA, suggesting that this protein could act as a DNA mimic. In a subsequent proteomic analysis, we found that the bacterial histone-like protein H-NS interacts with Arn, implying a new function. An electrophoretic mobility shift assay showed that Arn prevents H-NS from binding to the Escherichia coli hns and T4 p8.1 promoters. In vitro gene expression and electron microscopy analyses also indicated that Arn counteracts the gene-silencing effect of H-NS on a reporter gene. Because McrBC and H-NS both participate in the host defense system, our findings suggest that T4 Arn might knock down these mechanisms using its DNA mimicking properties. PMID:25118281

Structural determinants of HIV-1 nucleocapsid protein for cTAR DNA binding and destabilization, and correlation with inhibition of self-primed DNA synthesis.

PubMed

Beltz, Hervé; Clauss, Céline; Piémont, Etienne; Ficheux, Damien; Gorelick, Robert J; Roques, Bernard; Gabus, Caroline; Darlix, Jean-Luc; de Rocquigny, Hugues; Mély, Yves

2005-05-20

The nucleocapsid protein (NC) of human immunodeficiency virus type 1 (HIV-1) is formed of two highly conserved CCHC zinc fingers flanked by small basic domains. NC is required for the two obligatory strand transfers in viral DNA synthesis through its nucleic acid chaperoning properties. The first DNA strand transfer relies on NC's ability to bind and destabilize the secondary structure of complementary transactivation response region (cTAR) DNA, to inhibit self-priming, and to promote the annealing of cTAR to TAR RNA. To further investigate NC chaperone properties, our aim was to identify by fluorescence spectroscopy and gel electrophoresis, the NC structural determinants for cTAR binding and destabilization, and for the inhibition of self-primed DNA synthesis on a model system using a series of NC mutants and HIV-1 reverse transcriptase. NC destabilization and self-priming inhibition properties were found to be supported by the two fingers in their proper context and the basic (29)RAPRKKG(35) linker. The strict requirement of the native proximal finger suggests that its hydrophobic platform (Val13, Phe16, Thr24 and Ala25) is crucial for binding, destabilization and inhibition of self-priming. In contrast, only partial folding of the distal finger is required, probably for presenting the Trp37 residue in an appropriate orientation. Also, Trp37 and the hydrophobic residues of the proximal finger appear to be essential for the propagation of the melting from the cTAR ends up to the middle of the stem. Finally, both N-terminal and C-terminal basic domains contribute to cTAR binding but not to its destabilization.

DNA glycosylases search for and remove oxidized DNA bases.

PubMed

Wallace, Susan S

2013-12-01

This review article presents, an overview of the DNA glycosylases that recognize oxidized DNA bases using the Fpg/Nei family of DNA glycosylases as models for how structure can inform function. For example, even though human NEIL1 and the plant and fungal orthologs lack the zinc finger shown to be required for binding, DNA crystal structures revealed a "zincless finger" with the same properties. Moreover, the "lesion recognition loop" is not involved in lesion recognition, rather, it stabilizes 8-oxoG in the active site pocket. Unlike the other Fpg/Nei family members, Neil3 lacks two of the three void-filling residues that stabilize the DNA duplex and interact with the opposite strand to the damage which may account for its preference for lesions in single-stranded DNA. Also single-molecule approaches show that DNA glycosylases search for their substrates in a sea of undamaged DNA by using a wedge residue that is inserted into the DNA helix to probe for the presence of damage. Copyright © 2013 Wiley Periodicals, Inc.

Sequence-dependent DNA flexibility mediates DNase I cleavage.

PubMed

Heddi, Brahim; Abi-Ghanem, Josephine; Lavigne, Marc; Hartmann, Brigitte

2010-01-08

Understanding the preference of nonspecific proteins for certain DNA structural features requires an accurate description of the properties of free DNA, especially regarding their possible predisposition to adopt a conformation that favors the formation of a complex. Exploiting previous exhaustive NMR studies performed on free DNA oligomers, we investigated the molecular basis of DNase I sensitivity under conditions where DNase I binding limits the probability of cleavage. We showed that cleavage intensity was correlated with adjacent 3' phosphate linkage flexibility, monitored by (31)P chemical shifts. Examining NMR-refined DNA structures highlighted that sequence-dependent flexible phosphates were associated with large minor groove variations that may promote the affinity of DNase I, according to relevant DNA-protein complexes. In sum, this work demonstrates that specificity in DNA-DNase I interaction is mediated by DNA flexibility, which influences the induced-fit transitions required to form productive complexes.

Submolecular Structure and Orientation of Oligonucleotide Duplexes Tethered to Gold Electrodes Probed by Infrared Reflection Absorption Spectroscopy: Effect of the Electrode Potentials.

PubMed

Kékedy-Nagy, László; Ferapontova, Elena E; Brand, Izabella

2017-02-23

Unique electronic and ligand recognition properties of the DNA double helix provide basis for DNA applications in biomolecular electronic and biosensor devices. However, the relation between the structure of DNA at electrified interfaces and its electronic properties is still not well understood. Here, potential-driven changes in the submolecular structure of DNA double helices composed of either adenine-thymine (dAdT) 25 or cytosine-guanine (dGdC) 20 base pairs tethered to the gold electrodes are for the first time analyzed by in situ polarization modulation infrared reflection absorption spectroscopy (PM IRRAS) performed under the electrochemical control. It is shown that the conformation of the DNA duplexes tethered to gold electrodes via the C 6 alkanethiol linker strongly depends on the nucleic acid sequence composition. The tilt of purine and pyrimidine rings of the complementary base pairs (dAdT and dGdC) depends on the potential applied to the electrode. By contrast, neither the conformation nor orientation of the ionic in character phosphate-sugar backbone is affected by the electrode potentials. At potentials more positive than the potential of zero charge (pzc), a gradual tilting of the double helix is observed. In this tilted orientation, the planes of the complementary purine and pyrimidine rings lie ideally parallel to each other. These potentials do not affect the integral stability of the DNA double helix at the charged interface. At potentials more negative than the pzc, DNA helices adopt a vertical to the gold surface orientation. Tilt of the purine and pyrimidine rings depends on the composition of the double helix. In monolayers composed of (dAdT) 25 molecules the rings of the complementary base pairs lie parallel to each other. By contrast, the tilt of purine and pyrimidine rings in (dGdC) 20 helices depends on the potential applied to the electrode. Such potential-induced mobility of the complementary base pairs can destabilize the helix structure at a submolecular level. These pioneer results on the potential-driven changes in the submolecular structure of double stranded DNA adsorbed on conductive supports contribute to further understanding of the potential-driven sequence-specific electronic properties of surface-tethered oligonucleotides.

On the topology of chromatin fibres

PubMed Central

Barbi, Maria; Mozziconacci, Julien; Victor, Jean-Marc; Wong, Hua; Lavelle, Christophe

2012-01-01

The ability of cells to pack, use and duplicate DNA remains one of the most fascinating questions in biology. To understand DNA organization and dynamics, it is important to consider the physical and topological constraints acting on it. In the eukaryotic cell nucleus, DNA is organized by proteins acting as spools on which DNA can be wrapped. These proteins can subsequently interact and form a structure called the chromatin fibre. Using a simple geometric model, we propose a general method for computing topological properties (twist, writhe and linking number) of the DNA embedded in those fibres. The relevance of the method is reviewed through the analysis of magnetic tweezers single molecule experiments that revealed unexpected properties of the chromatin fibre. Possible biological implications of these results are discussed. PMID:24098838

Surface-immobilized DNAzyme-type biocatalysis

NASA Astrophysics Data System (ADS)

Stefan, Loic; Lavergne, Thomas; Spinelli, Nicolas; Defrancq, Eric; Monchaud, David

2014-02-01

The structure of the double helix of deoxyribonucleic acid (DNA, also called duplex-DNA) was elucidated sixty years ago by Watson, Crick, Wilkins and Franklin. Since then, DNA has continued to hold a fascination for researchers in diverse fields including medicine and nanobiotechnology. Nature has indeed excelled in diversifying the use of DNA: beyond its canonical role of repository of genetic information, DNA could also act as a nanofactory able to perform some complex catalytic tasks in an enzyme-mimicking manner. The catalytic capability of DNA was termed DNAzyme; in this context, a peculiar DNA structure, a quadruple helix also named quadruplex-DNA, has recently garnered considerable interest since its autonomous catalytic proficiency relies on its higher-order folding that makes it suitable to interact efficiently with hemin, a natural cofactor of many enzymes. Quadruplexes have thus been widely studied for their hemoprotein-like properties, chiefly peroxidase-like activity, i.e., their ability to perform hemin-mediated catalytic oxidation reactions. Recent literature is replete with applications of quadruplex-based peroxidase-mimicking DNAzyme systems. Herein, we take a further leap along the road to biochemical applications, assessing the actual efficiency of catalytic quadruplexes for the detection of picomolar levels of surface-bound analytes in an enzyme-linked immunosorbent (ELISA)-type assay. To this end, we exploit an innovative strategy based on the functionalization of DNA by a multitasking platform named RAFT (for regioselectivity addressable functionalized template), whose versatility enables the grafting of DNA whatever its nature (duplex-DNA, quadruplex-DNA, etc.). We demonstrate that the resulting biotinylated RAFT/quadruplex systems indeed acquire catalytic properties that allow for efficient luminescent detection of picomoles of surface-bound streptavidin. We also highlight some of the pitfalls that have to be faced during optimization, notably demonstrating that highly optimized experimental conditions can make DNA pre-catalysts catalytically competent whatever their secondary structures.

The Kinetics of the as Grown and Annealed Self-Assembled Monolayer Studied by Force Spectroscopy

NASA Astrophysics Data System (ADS)

Habib, Huma; Yasar, M.; Mehmood, S.; Rafique, Saima; Bhatti, A. S.; Naeem, Aisha

The growth of biological systems like DNA, peptides and proteins are accredited to the self-assembly processes from the molecular level to the nanoscale. The flawless immobilization of DNA on any surface is quite an important step to the development of DNA-based biosensors. The present paper reports the use of atomic force microscopy to determine the mechanical properties of the as grown and annealed self-assembled monolayer (SAM) as well as the mutated DNA immobilized on the SAM. The SAM of alkane thiol (16-mercapto-1-hexadecanol) was developed on Au surface, which was then annealed and analyzed for its structural and mechanical properties. The surface coverage, height and monolayer’s order was studied as a function of incubation time and annealing time. Excessive annealing led to the defragmentation and desorption of SAM structures due to breaking of hydrocarbon bonds. AFM was employed to determine the detach separation, pull-off and work of adhesion of the as grown and annealed SAM.

Internal-Modified Dithiol DNA-Directed Au Nanoassemblies: Geometrically Controlled Self-Assembly and Quantitative Surface-Enhanced Raman Scattering Properties

NASA Astrophysics Data System (ADS)

Yan, Yuan; Shan, Hangyong; Li, Min; Chen, Shu; Liu, Jianyu; Cheng, Yanfang; Ye, Cui; Yang, Zhilin; Lai, Xuandi; Hu, Jianqiang

2015-11-01

In this work, a hierarchical DNA-directed self-assembly strategy to construct structure-controlled Au nanoassemblies (NAs) has been demonstrated by conjugating Au nanoparticles (NPs) with internal-modified dithiol single-strand DNA (ssDNA) (Au-B-A or A-B-Au-B-A). It is found that the dithiol-ssDNA-modified Au NPs and molecule quantity of thiol-modified ssDNA grafted to Au NPs play critical roles in the assembly of geometrically controlled Au NAs. Through matching Au-DNA self-assembly units, geometrical structures of the Au NAs can be tailored from one-dimensional (1D) to quasi-2D and 2D. Au-B-A conjugates readily give 1D and quasi-2D Au NAs while 2D Au NAs can be formed by A-B-Au-B-A building blocks. Surface-enhanced Raman scattering (SERS) measurements and 3D finite-difference time domain (3D-FDTD) calculation results indicate that the geometrically controllable Au NAs have regular and linearly “hot spots”-number-depended SERS properties. For a certain number of NPs, the number of “hot spots” and accordingly enhancement factor of Au NAs can be quantitatively evaluated, which open a new avenue for quantitative analysis based on SERS technique.

Active microrheology of entangled biopolymer composites link polymer flexibility and length to molecular force response

NASA Astrophysics Data System (ADS)

Fitzpatrick, Robert; Hauer, Cole; Kyrillos, Carl; McGorty, Ryan; Robertson-Anderson, Rae

Entangled polymers have complex viscoelastic properties that are tuned by polymer lengths and flexibilities. Entangled composites of distinct polymers offer added versatility and display nonlinear mechanics, serving as a platform for multifunctional materials. To determine the role of flexibility and length in polymer composites we use optical tweezers and confocal microscopy to measure mechanical and structural properties of co-entangled actin and DNA. Actin filaments have lengths of 5-20 μm, comparable to their persistence length, while DNA of similar lengths have hundreds of persistence lengths per chain. To characterize the nonlinear mechanics of actin-DNA composites, we optically drive a microsphere through the composite and measure the induced force during and following strain. We characterize viscoelasticity and relaxation timescales; and determine the dependence of these quantities on the actin:DNA ratio (0:1-1:0) and DNA length (4-100 μm). We use confocal microscopy to image distinctly labeled co-entangled actin and DNA and characterize network homogeneity and fluctuations. Initial results show actin and DNA are well-integrated and form structurally homogenous networks that exhibit stiffness and relaxation times that increase nonlinearly with increased actin. NSF Career Award (DMR-1254340), AFOSR Young Investigator Program Award (FA95550-12-1-0315), Scialog Collaborative Innovation Award funed by Research Corp. for Scientific Advancement (24192).

Packaging DNA Origami into Viral Protein Cages.

PubMed

Linko, Veikko; Mikkilä, Joona; Kostiainen, Mauri A

2018-01-01

The DNA origami technique is a widely used method to create customized, complex, spatially well-defined two-dimensional (2D) and three-dimensional (3D) DNA nanostructures. These structures have huge potential to serve as smart drug-delivery vehicles and molecular devices in various nanomedical and biotechnological applications. However, so far only little is known about the behavior of these novel structures in living organisms or in cell culture/tissue models. Moreover, enhancing pharmacokinetic bioavailability and transfection properties of such structures still remains a challenge. One intriguing approach to overcome these issues is to coat DNA origami nanostructures with proteins or lipid membranes. Here, we show how cowpea chlorotic mottle virus (CCMV) capsid proteins (CPs) can be used for coating DNA origami nanostructures. We present a method for disassembling native CCMV particles and isolating the pure CP dimers, which can further bind and encapsulate a rectangular DNA origami shape. Owing to the highly programmable nature of DNA origami, packaging of DNA nanostructures into viral protein cages could find imminent uses in enhanced targeting and cellular delivery of various active nano-objects, such as enzymes and drug molecules.

[Molecular dynamics of immune complex of photoadduct-containing DNA with Fab-Anti-DNA antibody fragment].

PubMed

Akberova, N I; Zhmurov, A A; Nevzorova, T A; Litvinov, R I

2016-01-01

Antibodies to DNA play an important role in the pathogenesis of autoimmune diseases. The elucidation of structural mechanisms of both the antigen recognition and the interaction of anti-DNA antibodies with DNA will help to understand the role of DNA-containing immune complexes in various pathologies and can provide a basis for new treatment modalities. Moreover, the DNA-antibody complex is an analog of specific intracellular DNA-protein interactions. In this work, we used in silico molecular dynamic simulations of bimolecular complexes of the dsDNA segment containing the Fab fragment of an anti-DNA antibody to obtain the detailed thermodynamic and structural characteristics of dynamic intermolecular interactions. Using computationally modified crystal structure of the Fab-DNA complex (PDB ID: 3VW3), we studied the equilibrium molecular dynamics of the 64M-5 antibody Fab fragment associated with the dsDNA fragment containing the thymine dimer, the product of DNA photodamage. Amino acid residues that constitute paratopes and the complementary nucleotide epitopes for the Fab-DNA construct were identified. Stacking and electrostatic interactions were found to play the main role in mediating the most specific antibody-dsDNA contacts, while hydrogen bonds were less significant. These findings may shed light on the formation and properties of pathogenic anti-DNA antibodies in autoimmune diseases, such as systemic lupus erythematosus associated with skin photosensitivity and DNA photodamage.

Holliday Junction Thermodynamics and Structure: Coarse-Grained Simulations and Experiments

NASA Astrophysics Data System (ADS)

Wang, Wujie; Nocka, Laura M.; Wiemann, Brianne Z.; Hinckley, Daniel M.; Mukerji, Ishita; Starr, Francis W.

2016-03-01

Holliday junctions play a central role in genetic recombination, DNA repair and other cellular processes. We combine simulations and experiments to evaluate the ability of the 3SPN.2 model, a coarse-grained representation designed to mimic B-DNA, to predict the properties of DNA Holliday junctions. The model reproduces many experimentally determined aspects of junction structure and stability, including the temperature dependence of melting on salt concentration, the bias between open and stacked conformations, the relative populations of conformers at high salt concentration, and the inter-duplex angle (IDA) between arms. We also obtain a close correspondence between the junction structure evaluated by all-atom and coarse-grained simulations. We predict that, for salt concentrations at physiological and higher levels, the populations of the stacked conformers are independent of salt concentration, and directly observe proposed tetrahedral intermediate sub-states implicated in conformational transitions. Our findings demonstrate that the 3SPN.2 model captures junction properties that are inaccessible to all-atom studies, opening the possibility to simulate complex aspects of junction behavior.

DNA minicircles clarify the specific role of DNA structure on retroviral integration

PubMed Central

Pasi, Marco; Mornico, Damien; Volant, Stevenn; Juchet, Anna; Batisse, Julien; Bouchier, Christiane; Parissi, Vincent; Ruff, Marc; Lavery, Richard; Lavigne, Marc

2016-01-01

Chromatin regulates the selectivity of retroviral integration into the genome of infected cells. At the nucleosome level, both histones and DNA structure are involved in this regulation. We propose a strategy that allows to specifically study a single factor: the DNA distortion induced by the nucleosome. This strategy relies on mimicking this distortion using DNA minicircles (MCs) having a fixed rotational orientation of DNA curvature, coupled with atomic-resolution modeling. Contrasting MCs with linear DNA fragments having identical sequences enabled us to analyze the impact of DNA distortion on the efficiency and selectivity of integration. We observed a global enhancement of HIV-1 integration in MCs and an enrichment of integration sites in the outward-facing DNA major grooves. Both of these changes are favored by LEDGF/p75, revealing a new, histone-independent role of this integration cofactor. PFV integration is also enhanced in MCs, but is not associated with a periodic redistribution of integration sites, thus highlighting its distinct catalytic properties. MCs help to separate the roles of target DNA structure, histone modifications and integrase (IN) cofactors during retroviral integration and to reveal IN-specific regulation mechanisms. PMID:27439712

Structural characterization of a new lipid/DNA complex showing a selective transfection efficiency in ovarian cancer cells

NASA Astrophysics Data System (ADS)

Caracciolo, G.; Pozzi, D.; Caminiti, R.; Congiu Castellano, A.

2003-04-01

We investigated, for the first time, by using Energy Dispersive X-ray Diffraction, the structure of a new ternary cationic liposome formulated with dioleoyl trimethylammonium propane (DOTAP), 1,2-dioleoyl-3-phosphatidylethanolamine (DOPE) and cholesterol (Chol) (DDC) which has been recently found to have a selective high gene transfer ability in ovarian cancer cells. Our structural results provide a further experimental support to the widely accepted statement that there is not a simple and direct correlation between structure and transfection efficiency and that the factors controlling cationic lipid/DNA (CL-DNA) complexes-mediated gene transfer depend not only on the formulations of the cationic liposomes and their thermodynamic phase, but also significantly on the cell properties.

Structure-function relationships of new lipids designed for DNA transfection.

PubMed

Dittrich, Matthias; Heinze, Martin; Wölk, Christian; Funari, Sergio S; Dobner, Bodo; Möhwald, Helmuth; Brezesinski, Gerald

2011-08-22

Cationic liposome/DNA complexes can be used as nonviral vectors for direct delivery of DNA-based biopharmaceuticals to damaged cells and tissues. To obtain more effective and safer liposome-based gene transfection systems, two cationic lipids with identical head groups but different chain structures are investigated with respect to their in vitro gene-transfer activity, their cell-damaging characteristics, and their physicochemical properties. The gene-transfer activities of the two lipids are very different. Differential scanning calorimetry and synchrotron small- and wide-angle X-ray scattering give valuable structural insight. A subgel-like structure with high packing density and high phase-transition temperature from gel to liquid-crystalline state are found for lipid 7 (N'-2-[(2,6-diamino-1-oxohexyl)amino]ethyl-2,N-bis(hexadecyl)propanediamide) containing two saturated chains. Additionally, an ordered head-group lattice based on formation of a hydrogen-bond network is present. In contrast, lipid 8 (N'-2-[(2,6-diamino-1-oxohexyl)amino]ethyl-2-hexadecyl-N-[(9Z)-octadec-9-enyl]propanediamide) with one unsaturated and one saturated chain shows a lower phase-transition temperature and a reduced packing density. These properties enhance incorporation of the helper lipid cholesterol needed for gene transfection. Both lipids, either pure or in mixtures with cholesterol, form lamellar phases, which are preserved after addition of DNA. However, the system separates into phases containing DNA and phases without DNA. On increasing the temperature, DNA is released and only a lipid phase without intercalated DNA strands is observed. The conversion temperatures are very different in the two systems studied. The important parameter seems to be the charge density of the lipid membranes, which is a result of different solubility of cholesterol in the two lipid membranes. Therefore, different binding affinities of the DNA to the lipid mixtures are achieved. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Vibrational, structural and electronic properties investigation by DFT calculations and molecular docking studies with DNA topoisomerase II of strychnobrasiline type alkaloids: A theoretical approach for potentially bioactive molecules

NASA Astrophysics Data System (ADS)

Costa, Renyer A.; Oliveira, Kelson M. T.; Costa, Emmanoel Vilaça; Pinheiro, Maria L. B.

2017-10-01

A combined experimental and theoretical DFT study of the structural, vibrational and electronic properties of strychnobrasiline and 12-hydroxy-10,11-dimethoxystrychnobrasiline is presented using the Becke three-parameter Lee-Yang-Parr function (B3LYP) and 6-311G(2d,p) basis set. The theoretical geometry optimization data were compared with the X-ray data for a similar structure in the associated literature, showing close values. The calculated HOMO-LUMO gap values showed that the presence of substituents in the benzene ring influences the quantum properties which are directly related to the reactive properties. Theoretical UV spectra agreed well with the measured experimental data, with bands assigned. In addition, Natural Bond Orbitals (NBOs), Mapped molecular electrostatic potential surface (MEPS) and NLO calculations were also performed at the same theory level. The theoretical vibrational analysis revealed several characteristic vibrations that may be used as a diagnostic tool for other strychnobrasiline type alkaloids, simplifying their identification and structural characterization. Molecular docking calculations with DNA Topoisomerase II-DNA complex showed binding free energies values of -8.0 and -9.5 kcal/mol for strychnobrasiline and 12-hydroxy-10,11-dimethoxystrychnobrasiline respectively, while for amsacrine, used for the treatment of leukemia, the binding free energy ΔG presented a value of -10.0 kcal/mol, suggesting that strychnobrasiline derivative alkaloids might exhibit an antineoplastic activity.

An experimentally-informed coarse-grained 3-site-per-nucleotide model of DNA: Structure, thermodynamics, and dynamics of hybridization

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hinckley, Daniel M.; Freeman, Gordon S.; Whitmer, Jonathan K.

2013-10-14

A new 3-Site-Per-Nucleotide coarse-grained model for DNA is presented. The model includes anisotropic potentials between bases involved in base stacking and base pair interactions that enable the description of relevant structural properties, including the major and minor grooves. In an improvement over available coarse-grained models, the correct persistence length is recovered for both ssDNA and dsDNA, allowing for simulation of non-canonical structures such as hairpins. DNA melting temperatures, measured for duplexes and hairpins by integrating over free energy surfaces generated using metadynamics simulations, are shown to be in quantitative agreement with experiment for a variety of sequences and conditions. Hybridizationmore » rate constants, calculated using forward-flux sampling, are also shown to be in good agreement with experiment. The coarse-grained model presented here is suitable for use in biological and engineering applications, including nucleosome positioning and DNA-templated engineering.« less

A comparative approach for the investigation of biological information processing: An examination of the structure and function of computer hard drives and DNA

PubMed Central

2010-01-01

Background The robust storage, updating and utilization of information are necessary for the maintenance and perpetuation of dynamic systems. These systems can exist as constructs of metal-oxide semiconductors and silicon, as in a digital computer, or in the "wetware" of organic compounds, proteins and nucleic acids that make up biological organisms. We propose that there are essential functional properties of centralized information-processing systems; for digital computers these properties reside in the computer's hard drive, and for eukaryotic cells they are manifest in the DNA and associated structures. Methods Presented herein is a descriptive framework that compares DNA and its associated proteins and sub-nuclear structure with the structure and function of the computer hard drive. We identify four essential properties of information for a centralized storage and processing system: (1) orthogonal uniqueness, (2) low level formatting, (3) high level formatting and (4) translation of stored to usable form. The corresponding aspects of the DNA complex and a computer hard drive are categorized using this classification. This is intended to demonstrate a functional equivalence between the components of the two systems, and thus the systems themselves. Results Both the DNA complex and the computer hard drive contain components that fulfill the essential properties of a centralized information storage and processing system. The functional equivalence of these components provides insight into both the design process of engineered systems and the evolved solutions addressing similar system requirements. However, there are points where the comparison breaks down, particularly when there are externally imposed information-organizing structures on the computer hard drive. A specific example of this is the imposition of the File Allocation Table (FAT) during high level formatting of the computer hard drive and the subsequent loading of an operating system (OS). Biological systems do not have an external source for a map of their stored information or for an operational instruction set; rather, they must contain an organizational template conserved within their intra-nuclear architecture that "manipulates" the laws of chemistry and physics into a highly robust instruction set. We propose that the epigenetic structure of the intra-nuclear environment and the non-coding RNA may play the roles of a Biological File Allocation Table (BFAT) and biological operating system (Bio-OS) in eukaryotic cells. Conclusions The comparison of functional and structural characteristics of the DNA complex and the computer hard drive leads to a new descriptive paradigm that identifies the DNA as a dynamic storage system of biological information. This system is embodied in an autonomous operating system that inductively follows organizational structures, data hierarchy and executable operations that are well understood in the computer science industry. Characterizing the "DNA hard drive" in this fashion can lead to insights arising from discrepancies in the descriptive framework, particularly with respect to positing the role of epigenetic processes in an information-processing context. Further expansions arising from this comparison include the view of cells as parallel computing machines and a new approach towards characterizing cellular control systems. PMID:20092652

A comparative approach for the investigation of biological information processing: an examination of the structure and function of computer hard drives and DNA.

PubMed

D'Onofrio, David J; An, Gary

2010-01-21

The robust storage, updating and utilization of information are necessary for the maintenance and perpetuation of dynamic systems. These systems can exist as constructs of metal-oxide semiconductors and silicon, as in a digital computer, or in the "wetware" of organic compounds, proteins and nucleic acids that make up biological organisms. We propose that there are essential functional properties of centralized information-processing systems; for digital computers these properties reside in the computer's hard drive, and for eukaryotic cells they are manifest in the DNA and associated structures. Presented herein is a descriptive framework that compares DNA and its associated proteins and sub-nuclear structure with the structure and function of the computer hard drive. We identify four essential properties of information for a centralized storage and processing system: (1) orthogonal uniqueness, (2) low level formatting, (3) high level formatting and (4) translation of stored to usable form. The corresponding aspects of the DNA complex and a computer hard drive are categorized using this classification. This is intended to demonstrate a functional equivalence between the components of the two systems, and thus the systems themselves. Both the DNA complex and the computer hard drive contain components that fulfill the essential properties of a centralized information storage and processing system. The functional equivalence of these components provides insight into both the design process of engineered systems and the evolved solutions addressing similar system requirements. However, there are points where the comparison breaks down, particularly when there are externally imposed information-organizing structures on the computer hard drive. A specific example of this is the imposition of the File Allocation Table (FAT) during high level formatting of the computer hard drive and the subsequent loading of an operating system (OS). Biological systems do not have an external source for a map of their stored information or for an operational instruction set; rather, they must contain an organizational template conserved within their intra-nuclear architecture that "manipulates" the laws of chemistry and physics into a highly robust instruction set. We propose that the epigenetic structure of the intra-nuclear environment and the non-coding RNA may play the roles of a Biological File Allocation Table (BFAT) and biological operating system (Bio-OS) in eukaryotic cells. The comparison of functional and structural characteristics of the DNA complex and the computer hard drive leads to a new descriptive paradigm that identifies the DNA as a dynamic storage system of biological information. This system is embodied in an autonomous operating system that inductively follows organizational structures, data hierarchy and executable operations that are well understood in the computer science industry. Characterizing the "DNA hard drive" in this fashion can lead to insights arising from discrepancies in the descriptive framework, particularly with respect to positing the role of epigenetic processes in an information-processing context. Further expansions arising from this comparison include the view of cells as parallel computing machines and a new approach towards characterizing cellular control systems.

Simultaneous Drug Targeting of the Promoter MYC G-Quadruplex and BCL2 i-Motif in Diffuse Large B-Cell Lymphoma Delays Tumor Growth.

PubMed

Kendrick, Samantha; Muranyi, Andrea; Gokhale, Vijay; Hurley, Laurence H; Rimsza, Lisa M

2017-08-10

Secondary DNA structures are uniquely poised as therapeutic targets due to their molecular switch function in turning gene expression on or off and scaffold-like properties for protein and small molecule interaction. Strategies to alter gene transcription through these structures thus far involve targeting single DNA conformations. Here we investigate the feasibility of simultaneously targeting different secondary DNA structures to modulate two key oncogenes, cellular-myelocytomatosis (MYC) and B-cell lymphoma gene-2 (BCL2), in diffuse large B-cell lymphoma (DLBCL). Cotreatment with previously identified ellipticine and pregnanol derivatives that recognize the MYC G-quadruplex and BCL2 i-motif promoter DNA structures lowered mRNA levels and subsequently enhanced sensitivity to a standard chemotherapy drug, cyclophosphamide, in DLBCL cell lines. In vivo repression of MYC and BCL2 in combination with cyclophosphamide also significantly slowed tumor growth in DLBCL xenograft mice. Our findings demonstrate concurrent targeting of different DNA secondary structures offers an effective, precise, medicine-based approach to directly impede transcription and overcome aberrant pathways in aggressive malignancies.

Nucleosome core particles containing a poly(dA.dT) sequence element exhibit a locally distorted DNA structure.

PubMed

Bao, Yunhe; White, Cindy L; Luger, Karolin

2006-08-25

Poly(dA.dT) DNA sequence elements are thought to promote transcription by either excluding nucleosomes or by altering their structural or dynamic properties. Here, the stability and structure of a defined nucleosome core particle containing a 16 base-pair poly(dA.dT) element (A16 NCP) was investigated. The A16 NCP requires a significantly higher temperature for histone octamer sliding in vitro compared to comparable nucleosomes that do not contain a poly(dA.dT) element. Fluorescence resonance energy transfer showed that the interactions between the nucleosomal DNA ends and the histone octamer were destabilized in A16 NCP. The crystal structure of A16 NCP was determined to a resolution of 3.2 A. The overall structure was maintained except for local deviations in DNA conformation. These results are consistent with previous in vivo and in vitro observations that poly(dA.dT) elements cause only modest changes in DNA accessibility and modest increases in steady-state transcription levels.

Extended Fluorescent Resonant Energy Transfer in DNA Constructs

NASA Astrophysics Data System (ADS)

Oh, Taeseok

This study investigates the use of surfactants and metal cations for the enhancement of long range fluorescent resonant energy transfer (FRET) and the antenna effect in DNA structures with multiple fluorescent dyes. Double-stranded (ds) DNA structures were formed by hybridization of 21mer DNA oligonucleotides with different arrangements of three fluorescent TAMRA donor dyes with two different complementary 21mer oligonucleotides with one fluorescent TexasRed acceptor dye. In such DNA structures, hydrophobic interactions between the fluorescent dyes in close proximity produces dimerization which along with other quenching mechanisms leads to significant reduction of fluorescent emission properties. Addition of the surfactants Triton X-100, cetyltrimethyl ammonium bromide (CTAB) and sodium dodecyl sulfate (SDS) along with sodium cations (Na+) and divalent magnesium cations (Mg 2+) were tested for their ability to reduce quenching of the fluorescent dyes and improve overall fluorescent emission, the long range FRET and the antenna effect properties. When the neutral (uncharged) surfactant Triton X-100 was added to the FRET ds-DNA hybrid structures with three TAMRA donors and one TexasRed acceptor, dye dimerization and emission quenching remained unaffected. However, for the positively charged CTAB surfactant at concentrations of 100 uM or higher, the neutralization of the negatively charged ds-DNA backbone by the cationic surfactant micelles was found to reduce TAMRA dye dimerization and emission quenching and improve TexasRed quantum yield, resulting in much higher FRET efficiencies and an enhanced antenna effect. This improvement is likely due to the CTAB molecules covering or sheathing the fluorescent donor and acceptor dyes which breaks up the dimerized dye complexes and prevents further quenching from interactions with water molecules and guanine bases in the DNA structure. While the negatively charged SDS surfactant alone was not able to reduce dimerization and emission quenching due to repulsive forces between DNA and SDS micelles, the addition of cations such as sodium ions (Na+) and divalent magnesium ions (Mg2+) did lead to a significant reduction in the dimerization and emission quenching resulting in much higher FRET efficiency and an enhanced antenna effect. It appears that when the repulsive electrostatic forces are screened by the cations (Mg2+ in particular), the SDS micelles can approach the FRET ds-DNA structures thereby sheathing or insulating the TAMRA and TexasRed dyes. Overall, the study provides a viable strategy for using combinations of surfactants and cations to reduce adverse fluorescent dye and other quenching mechanisms and improve the overall long distance FRET efficiency and the antenna effect in DNA structures with multi-donor and single acceptor fluorescent dye groups.

Construction of a fuzzy and Boolean logic gates based on DNA.

PubMed

Zadegan, Reza M; Jepsen, Mette D E; Hildebrandt, Lasse L; Birkedal, Victoria; Kjems, Jørgen

2015-04-17

Logic gates are devices that can perform logical operations by transforming a set of inputs into a predictable single detectable output. The hybridization properties, structure, and function of nucleic acids can be used to make DNA-based logic gates. These devices are important modules in molecular computing and biosensing. The ideal logic gate system should provide a wide selection of logical operations, and be integrable in multiple copies into more complex structures. Here we show the successful construction of a small DNA-based logic gate complex that produces fluorescent outputs corresponding to the operation of the six Boolean logic gates AND, NAND, OR, NOR, XOR, and XNOR. The logic gate complex is shown to work also when implemented in a three-dimensional DNA origami box structure, where it controlled the position of the lid in a closed or open position. Implementation of multiple microRNA sensitive DNA locks on one DNA origami box structure enabled fuzzy logical operation that allows biosensing of complex molecular signals. Integrating logic gates with DNA origami systems opens a vast avenue to applications in the fields of nanomedicine for diagnostics and therapeutics. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Interrelations of secondary structure stability and DNA-binding affinity in the bacteriophage SPO1-encoded type II DNA-binding protein TF1.

PubMed

Andera, L; Spangler, C J; Galeone, A; Mayol, L; Geiduschek, E P

1994-02-11

TF1, a homodimeric DNA-binding and -bending protein with a preference for hydroxymethyluracil-containing DNA is the Bacillus subtilis-encoded homolog of the bacterial HU proteins and of the E. coli integration host factor. A temperature-sensitive mutation at amino acid 25 of TF1 (L25-->A) and two intragenic second site revertants at amino acids 15 (E15-->G) and 32 (L32-->I) were previously identified and their effects on virus development were examined. The DNA-binding properties of these proteins and the thermal stability of their secondary structures have now been analyzed. Amino acids 15 and 32 are far removed from the putative DNA-binding domains of TF1 but changes there exert striking effects on DNA affinity that correlate with effects on structure. The double mutant protein TF1-G15I32 binds to a preferred site in hydroxymethyluracil-containing DNA 40 times more tightly, denatures at higher temperature (delta tm = 21 degrees C), and also exchanges subunits much more slowly than does the wild-type protein. The L25-->A mutation makes TF1 secondary structure and DNA-binding highly salt concentration-dependent. The E15-->G mutation partly suppresses this effect: secondary structure of TF1-A25G15 is restored at 21 degrees C by 1 M NaCl or, at low NaCl concentration, by binding to DNA.

Mutations altering the cleavage specificity of a homing endonuclease

PubMed Central

Seligman, Lenny M.; Chisholm, Karen M.; Chevalier, Brett S.; Chadsey, Meggen S.; Edwards, Samuel T.; Savage, Jeremiah H.; Veillet, Adeline L.

2002-01-01

The homing endonuclease I-CreI recognizes and cleaves a particular 22 bp DNA sequence. The crystal structure of I-CreI bound to homing site DNA has previously been determined, leading to a number of predictions about specific protein–DNA contacts. We test these predictions by analyzing a set of endonuclease mutants and a complementary set of homing site mutants. We find evidence that all structurally predicted I-CreI/DNA contacts contribute to DNA recognition and show that these contacts differ greatly in terms of their relative importance. We also describe the isolation of a collection of altered specificity I-CreI derivatives. The in vitro DNA-binding and cleavage properties of two such endonucleases demonstrate that our genetic approach is effective in identifying homing endonucleases that recognize and cleave novel target sequences. PMID:12202772

An asymmetric structure of the Bacillus subtilis replication terminator protein in complex with DNA.

PubMed

Vivian, J P; Porter, C J; Wilce, J A; Wilce, M C J

2007-07-13

In Bacillus subtilis, the termination of DNA replication via polar fork arrest is effected by a specific protein:DNA complex formed between the replication terminator protein (RTP) and DNA terminator sites. We report the crystal structure of a replication terminator protein homologue (RTP.C110S) of B. subtilis in complex with the high affinity component of one of its cognate DNA termination sites, known as the TerI B-site, refined at 2.5 A resolution. The 21 bp RTP:DNA complex displays marked structural asymmetry in both the homodimeric protein and the DNA. This is in contrast to the previously reported complex formed with a symmetrical TerI B-site homologue. The induced asymmetry is consistent with the complex's solution properties as determined using NMR spectroscopy. Concomitant with this asymmetry is variation in the protein:DNA binding pattern for each of the subunits of the RTP homodimer. It is proposed that the asymmetric "wing" positions, as well as other asymmetrical features of the RTP:DNA complex, are critical for the cooperative binding that underlies the mechanism of polar fork arrest at the complete terminator site.

Dynamics of water around the complex structures formed between the KH domains of far upstream element binding protein and single-stranded DNA molecules

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chakraborty, Kaushik; Bandyopadhyay, Sanjoy, E-mail: sanjoy@chem.iitkgp.ernet.in

2015-07-28

Single-stranded DNA (ss-DNA) binding proteins specifically bind to the single-stranded regions of the DNA and protect it from premature annealing, thereby stabilizing the DNA structure. We have carried out atomistic molecular dynamics simulations of the aqueous solutions of two DNA binding K homology (KH) domains (KH3 and KH4) of the far upstream element binding protein complexed with two short ss-DNA segments. Attempts have been made to explore the influence of the formation of such complex structures on the microscopic dynamics and hydrogen bond properties of the interfacial water molecules. It is found that the water molecules involved in bridging themore » ss-DNA segments and the protein domains form a highly constrained thin layer with extremely retarded mobility. These water molecules play important roles in freezing the conformational oscillations of the ss-DNA oligomers and thereby forming rigid complex structures. Further, it is demonstrated that the effect of complexation on the slow long-time relaxations of hydrogen bonds at the interface is correlated with hindered motions of the surrounding water molecules. Importantly, it is observed that the highly restricted motions of the water molecules bridging the protein and the DNA components in the complexed forms originate from more frequent hydrogen bond reformations.« less

Altering DNA-Programmable Colloidal Crystallization Paths by Modulating Particle Repulsion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Mary X.; Brodin, Jeffrey D.; Millan, Jaime A.

Colloidal crystal engineering with DNA can be used to realize precise control over nanoparticle (NP) arrangement. Here, we investigate a case of DNA-based assembly where the properties of DNA as a polyelectrolyte brush are employed to alter a hybridization-driven NP crystallization pathway. Using the co-assembly of DNA-conjugated proteins and spherical gold 2 nanoparticles (AuNPs) as a model system, we explore how steric repulsion between non-complementary, neighboring DNA-NPs due to overlapping DNA shells can influence their ligand-directed behavior. Specifically, our experimental data coupled with coarse-grained molecular dynamics (MD) simulations reveal that by changing factors related to NP repulsion, two structurally distinctmore » outcomes can be achieved. When steric repulsion between DNA-AuNPs is significantly greater than that between DNA-proteins, a lower packing density crystal lattice is favored over the structure that is predicted by design rules based on DNA-hybridization considerations alone. This is enabled by the large difference in DNA density on AuNPs versus proteins and can be tuned by modulating the flexibility, and thus conformational entropy, of the DNA on the constituent particles. At intermediate ligand flexibility, the crystallization pathways are energetically similar and the structural outcome can be adjusted using the density of DNA duplexes on DNA-AuNPs and by screening the Coulomb potential between them. Such lattices are shown to undergo dynamic reorganization upon changing salt concentration. These data help elucidate the structural considerations necessary for understanding repulsive forces in DNA-assembly and lay the groundwork for using them to increase architectural diversity in engineering colloidal crystals.« less

Unifying the DNA End-processing Roles of the Artemis Nuclease

PubMed Central

Chang, Howard H. Y.; Watanabe, Go; Lieber, Michael R.

2015-01-01

Artemis is a member of the metallo-β-lactamase protein family of nucleases. It is essential in vertebrates because, during V(D)J recombination, the RAG complex generates hairpins when it creates the double strand breaks at V, D, and J segments, and Artemis is required to open the hairpins so that they can be joined. Artemis is a diverse endo- and exonuclease, and creating a unified model for its wide range of nuclease properties has been challenging. Here we show that Artemis resects iteratively into blunt DNA ends with an efficiency that reflects the AT-richness of the DNA end. GC-rich ends are not cut by Artemis alone because of a requirement for DNA end breathing (and confirmed using fixed pseudo-Y structures). All DNA ends are cut when both the DNA-dependent protein kinase catalytic subunit and Ku accompany Artemis but not when Ku is omitted. These are the first biochemical data demonstrating a Ku dependence of Artemis action on DNA ends of any configuration. The action of Artemis at blunt DNA ends is slower than at overhangs, consistent with a requirement for a slow DNA end breathing step preceding the cut. The AT sequence dependence, the order of strand cutting, the length of the cuts, and the Ku-dependence of Artemis action at blunt ends can be reconciled with the other nucleolytic properties of both Artemis and Artemis·DNA-PKcs in a model incorporating DNA end breathing of blunt ends to form transient single to double strand boundaries that have structural similarities to hairpins and fixed 5′ and 3′ overhangs. PMID:26276388

Topology of zigzag chromatin.

PubMed

Strogatz, S

1983-08-21

An enormous length of DNA is packaged in the nuclei of eukaryotic cells. This is achieved through several intermediate levels of compaction, ranging from the double helix to the chromosome. The nucleosome is now firmly established as the first level of chromatin structure. Next it appears that the nucleosomes are themselves stacked in a two-track array, with a dinucleosome repeat. Several winding patterns of DNA are compatible with such a structure. It is shown here that, compared to other feasible DNA paths, the observed winding pattern has remarkable topological properties. The possible biological significance of this peculiarity is discussed.

Relative information content of polymorphic microsatellites and mitochondrial DNA for inferring dispersal and population genetic structure in the olive sea snake, Aipysurus laevis.

PubMed

Lukoschek, V; Waycott, M; Keogh, J S

2008-07-01

Polymorphic microsatellites are widely considered more powerful for resolving population structure than mitochondrial DNA (mtDNA) markers, particularly for recently diverged lineages or geographically proximate populations. Weaker population subdivision for biparentally inherited nuclear markers than maternally inherited mtDNA may signal male-biased dispersal but can also be attributed to marker-specific evolutionary characteristics and sampling properties. We discriminated between these competing explanations with a population genetic study on olive sea snakes, Aipysurus laevis. A previous mtDNA study revealed strong regional population structure for A. laevis around northern Australia, where Pleistocene sea-level fluctuations have influenced the genetic signatures of shallow-water marine species. Divergences among phylogroups dated to the Late Pleistocene, suggesting recent range expansions by previously isolated matrilines. Fine-scale population structure within regions was, however, poorly resolved for mtDNA. In order to improve estimates of fine-scale genetic divergence and to compare population structure between nuclear and mtDNA, 354 olive sea snakes (previously sequenced for mtDNA) were genotyped for five microsatellite loci. F statistics and Bayesian multilocus genotype clustering analyses found similar regional population structure as mtDNA and, after standardizing microsatellite F statistics for high heterozygosities, regional divergence estimates were quantitatively congruent between marker classes. Over small spatial scales, however, microsatellites recovered almost no genetic structure and standardized F statistics were orders of magnitude smaller than for mtDNA. Three tests for male-biased dispersal were not significant, suggesting that recent demographic expansions to the typically large population sizes of A. laevis have prevented microsatellites from reaching mutation-drift equilibrium and local populations may still be diverging.

Polyplex formation between four-arm poly(ethylene oxide)-b-poly(2-(diethylamino)ethyl methacrylate) and plasmid DNA in gene delivery.

PubMed

He, E; Yue, C Y; Simeon, F; Zhou, L H; Too, H P; Tam, K C

2009-12-01

Amphiphilic polyelectrolytes comprising cationic and uncharged hydrophilic segments condensed negatively charged DNA to form a core-shell structure stabilized by a layer of hydrophilic corona chains. At physiological pH, four-arm star-shaped poly(ethylene oxide)-b-poly(2-(diethylamino)ethyl methacrylate) (four-arm PEO-b-PDEAEMA) block copolymer possessed positively charged amine groups that interacted with negatively charged plasmid DNA to form polymer/DNA complexes. The mechanism and physicochemical properties of the complex formation were investigated at varying molar ratio of amine groups on polymer chains and phosphate group on plasmid DNA segments (N/P ratio). The capability of the star block copolymer to condense DNA was demonstrated through gel electrophoresis and ethidium bromide exclusion assay. In the absence of salt, the hydrodynamic radius of polyplexes was about 94 nm at low polymer/DNA ratio, and it decreased to about 34 nm at large N/P ratios, forming a compact spherical structure with a weighted average molecular weight of 4.39 +/- 0.22 x 10(6) g/mol. Approximately 15 polymeric chains were required to condense a plasmid DNA. The addition of monovalent salt to the polyplexes significantly altered the size of the complexes, which would have an impact on cell transfection. Because of the electrostatic interaction induced by the diffusion of small ions, the polyplex increased in size to about 53 nm with a less compact structure. In vitro cytotoxicty of polymer and polymer/pDNA complexes were evaluated, and the polyplexes exhibited low toxicity at low N/P ratios. At N/P ratio of 4.5, the four-arm PEO-b-PDEAEMA showed the highest level of transfection in Neuro-2A cells. These observations showed that the star-shaped multi-arm polymers offers interesting properties in self-association and condensation ability for plasmid DNA and can serve as a nonviral DNA delivery system. Copyright 2008 Wiley Periodicals, Inc.

Lesion search and recognition by thymine DNA glycosylase revealed by single molecule imaging

PubMed Central

Buechner, Claudia N.; Maiti, Atanu; Drohat, Alexander C.; Tessmer, Ingrid

2015-01-01

The ability of DNA glycosylases to rapidly and efficiently detect lesions among a vast excess of nondamaged DNA bases is vitally important in base excision repair (BER). Here, we use single molecule imaging by atomic force microscopy (AFM) supported by a 2-aminopurine fluorescence base flipping assay to study damage search by human thymine DNA glycosylase (hTDG), which initiates BER of mutagenic and cytotoxic G:T and G:U mispairs in DNA. Our data reveal an equilibrium between two conformational states of hTDG–DNA complexes, assigned as search complex (SC) and interrogation complex (IC), both at target lesions and undamaged DNA sites. Notably, for both hTDG and a second glycosylase, hOGG1, which recognizes structurally different 8-oxoguanine lesions, the conformation of the DNA in the SC mirrors innate structural properties of their respective target sites. In the IC, the DNA is sharply bent, as seen in crystal structures of hTDG lesion recognition complexes, which likely supports the base flipping required for lesion identification. Our results support a potentially general concept of sculpting of glycosylases to their targets, allowing them to exploit the energetic cost of DNA bending for initial lesion sensing, coupled with continuous (extrahelical) base interrogation during lesion search by DNA glycosylases. PMID:25712093

Structurally Ordered Nanowire Formation from Co-Assembly of DNA Origami and Collagen-Mimetic Peptides

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jiang, Tao; Meyer, Travis A.; Modlin, Charles

In this paper, we describe the co-assembly of two different building units: collagen-mimetic peptides and DNA origami. Two peptides CP ++ and sCP ++ are designed with a sequence comprising a central block (Pro-Hyp-Gly) and two positively charged domains (Pro-Arg-Gly) at both N- and C-termini. Co-assembly of peptides and DNA origami two-layer (TL) nanosheets affords the formation of one-dimensional nanowires with repeating periodicity of similar to 10 nm. Structural analyses suggest a face-to-face stacking of DNA nanosheets with peptides aligned perpendicularly to the sheet surfaces. We demonstrate the potential of selective peptide-DNA association between face-to-face and edge-to-edge packing by tailoringmore » the size of DNA nanostructures. Finally, this study presents an attractive strategy to create hybrid biomolecular assemblies from peptide and DNA-based building blocks that takes advantage of the intrinsic chemical and physical properties of the respective components to encode structural and, potentially, functional complexity within readily accessible biomimetic materials.« less

Structurally Ordered Nanowire Formation from Co-Assembly of DNA Origami and Collagen-Mimetic Peptides

DOE PAGES

Jiang, Tao; Meyer, Travis A.; Modlin, Charles; ...

2017-09-26

In this paper, we describe the co-assembly of two different building units: collagen-mimetic peptides and DNA origami. Two peptides CP ++ and sCP ++ are designed with a sequence comprising a central block (Pro-Hyp-Gly) and two positively charged domains (Pro-Arg-Gly) at both N- and C-termini. Co-assembly of peptides and DNA origami two-layer (TL) nanosheets affords the formation of one-dimensional nanowires with repeating periodicity of similar to 10 nm. Structural analyses suggest a face-to-face stacking of DNA nanosheets with peptides aligned perpendicularly to the sheet surfaces. We demonstrate the potential of selective peptide-DNA association between face-to-face and edge-to-edge packing by tailoringmore » the size of DNA nanostructures. Finally, this study presents an attractive strategy to create hybrid biomolecular assemblies from peptide and DNA-based building blocks that takes advantage of the intrinsic chemical and physical properties of the respective components to encode structural and, potentially, functional complexity within readily accessible biomimetic materials.« less

Biochemical studies of the Saccharomyces cerevisiae Mph1 helicase on junction-containing DNA structures

PubMed Central

Kang, Young-Hoon; Munashingha, Palinda Ruvan; Lee, Chul-Hwan; Nguyen, Tuan Anh; Seo, Yeon-Soo

2012-01-01

Saccharomyces cerevisiae Mph1 is a 3–5′ DNA helicase, required for the maintenance of genome integrity. In order to understand the ATPase/helicase role of Mph1 in genome stability, we characterized its helicase activity with a variety of DNA substrates, focusing on its action on junction structures containing three or four DNA strands. Consistent with its 3′ to 5′ directionality, Mph1 displaced 3′-flap substrates in double-fixed or equilibrating flap substrates. Surprisingly, Mph1 displaced the 5′-flap strand more efficiently than the 3′ flap strand from double-flap substrates, which is not expected for a 3–5′ DNA helicase. For this to occur, Mph1 required a threshold size (>5 nt) of 5′ single-stranded DNA flap. Based on the unique substrate requirements of Mph1 defined in this study, we propose that the helicase/ATPase activity of Mph1 play roles in converting multiple-stranded DNA structures into structures cleavable by processing enzymes such as Fen1. We also found that the helicase activity of Mph1 was used to cause structural alterations required for restoration of replication forks stalled due to damaged template. The helicase properties of Mph1 reported here could explain how it resolves D-loop structure, and are in keeping with a model proposed for the error-free damage avoidance pathway. PMID:22090425

Structure and dynamics of double helical DNA in torsion angle hyperspace: a molecular mechanics approach.

PubMed

Borkar, Aditi; Ghosh, Indira; Bhattacharyya, Dhananjay

2010-04-01

Analysis of the conformational space populated by the torsion angles and the correlation between the conformational energy and the sequence of DNA are important for fully understanding DNA structure and function. Presence of seven variable torsion angles about single covalent bonds in DNA main chain puts a big challenge for such analysis. We have carried out restrained energy minimization studies for four representative dinucleosides, namely d(ApA):d(TpT), d(CpG):d(CpG), d(GpC):d(GpC) and d(CpA):d(TpG) to determine the energy hyperspace of DNA in context to the values of the torsion angles and the structural properties of the DNA conformations populating the favorable regions of this energy hyperspace. The torsion angles were manipulated by constraining their values at the reference points and then performing energy minimization. The energy minima obtained on the potential energy contour plots mostly correspond to the conformations populated in crystal structures of DNA. Some novel favorable conformations that are not present in crystal structure data are also found. The plots also suggest few low energy routes for conformational transitions or the associated energy barrier heights. Analyses of base pairing and stacking possibility reveal structural changes accompanying these transitions as well as the flexibility of different base steps towards variations in different torsion angles.

Repair-Resistant DNA Lesions

PubMed Central

2017-01-01

The eukaryotic global genomic nucleotide excision repair (GG-NER) pathway is the major mechanism that removes most bulky and some nonbulky lesions from cellular DNA. There is growing evidence that certain DNA lesions are repaired slowly or are entirely resistant to repair in cells, tissues, and in cell extract model assay systems. It is well established that the eukaryotic DNA lesion-sensing proteins do not detect the damaged nucleotide, but recognize the distortions/destabilizations in the native DNA structure caused by the damaged nucleotides. In this article, the nature of the structural features of certain bulky DNA lesions that render them resistant to NER, or cause them to be repaired slowly, is compared to that of those that are good-to-excellent NER substrates. Understanding the structural features that distinguish NER-resistant DNA lesions from good NER substrates may be useful for interpreting the biological significance of biomarkers of exposure of human populations to genotoxic environmental chemicals. NER-resistant lesions can survive to replication and cause mutations that can initiate cancer and other diseases. Furthermore, NER diminishes the efficacy of certain chemotherapeutic drugs, and the design of more potent pharmaceuticals that resist repair can be advanced through a better understanding of the structural properties of DNA lesions that engender repair-resistance. PMID:28750166

Imaging chromatin nanostructure with binding-activated localization microscopy based on DNA structure fluctuations

PubMed Central

Szczurek, Aleksander; Klewes, Ludger; Xing, Jun; Gourram, Amine; Birk, Udo; Knecht, Hans; Dobrucki, Jurek W.; Mai, Sabine

2017-01-01

Abstract Advanced light microscopy is an important tool for nanostructure analysis of chromatin. In this report we present a general concept for Single Molecule localization Microscopy (SMLM) super-resolved imaging of DNA-binding dyes based on modifying the properties of DNA and the dye. By careful adjustment of the chemical environment leading to local, reversible DNA melting and hybridization control over the fluorescence signal of the DNA-binding dye molecules can be introduced. We postulate a transient binding as the basis for our variation of binding-activated localization microscopy (BALM). We demonstrate that several intercalating and minor-groove binding DNA dyes can be used to register (optically isolate) only a few DNA-binding dye signals at a time. To highlight this DNA structure fluctuation-assisted BALM (fBALM), we applied it to measure, for the first time, nanoscale differences in nuclear architecture in model ischemia with an anticipated structural resolution of approximately 50 nm. Our data suggest that this approach may open an avenue for the enhanced microscopic analysis of chromatin nano-architecture and hence the microscopic analysis of nuclear structure aberrations occurring in various pathological conditions. It may also become possible to analyse nuclear nanostructure differences in different cell types, stages of development or environmental stress conditions. PMID:28082388

DNA recognition by synthetic constructs.

PubMed

Pazos, Elena; Mosquera, Jesús; Vázquez, M Eugenio; Mascareñas, José L

2011-09-05

The interaction of transcription factors with specific DNA sites is key for the regulation of gene expression. Despite the availability of a large body of structural data on protein-DNA complexes, we are still far from fully understanding the molecular and biophysical bases underlying such interactions. Therefore, the development of non-natural agents that can reproduce the DNA-recognition properties of natural transcription factors remains a major and challenging goal in chemical biology. In this review we summarize the basics of double-stranded DNA recognition by transcription factors, and describe recent developments in the design and preparation of synthetic DNA binders. We mainly focus on synthetic peptides that have been designed by following the DNA interaction of natural proteins, and we discuss how the tools of organic synthesis can be used to make artificial constructs equipped with functionalities that introduce additional properties to the recognition process, such as sensing and controllability. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Inferring coarse-grain histone-DNA interaction potentials from high-resolution structures of the nucleosome

NASA Astrophysics Data System (ADS)

Meyer, Sam; Everaers, Ralf

2015-02-01

The histone-DNA interaction in the nucleosome is a fundamental mechanism of genomic compaction and regulation, which remains largely unknown despite increasing structural knowledge of the complex. In this paper, we propose a framework for the extraction of a nanoscale histone-DNA force-field from a collection of high-resolution structures, which may be adapted to a larger class of protein-DNA complexes. We applied the procedure to a large crystallographic database extended by snapshots from molecular dynamics simulations. The comparison of the structural models first shows that, at histone-DNA contact sites, the DNA base-pairs are shifted outwards locally, consistent with locally repulsive forces exerted by the histones. The second step shows that the various force profiles of the structures under analysis derive locally from a unique, sequence-independent, quadratic repulsive force-field, while the sequence preferences are entirely due to internal DNA mechanics. We have thus obtained the first knowledge-derived nanoscale interaction potential for histone-DNA in the nucleosome. The conformations obtained by relaxation of nucleosomal DNA with high-affinity sequences in this potential accurately reproduce the experimental values of binding preferences. Finally we address the more generic binding mechanisms relevant to the 80% genomic sequences incorporated in nucleosomes, by computing the conformation of nucleosomal DNA with sequence-averaged properties. This conformation differs from those found in crystals, and the analysis suggests that repulsive histone forces are related to local stretch tension in nucleosomal DNA, mostly between adjacent contact points. This tension could play a role in the stability of the complex.

Defect tolerance and the effect of structural inhomogeneity in plasmonic DNA-nanoparticle superlattices

DOE PAGES

Ross, Michael B.; Ku, Jessie C.; Blaber, Martin G.; ...

2015-08-03

Bottom-up assemblies of plasmonic nanoparticles exhibit unique optical effects such as tunable reflection, optical cavity modes, and tunable photonic resonances. In this paper, we compare detailed simulations with experiment to explore the effect of structural inhomogeneity on the optical response in DNA-gold nanoparticle superlattices. In particular, we explore the effect of background environment, nanoparticle polydispersity (>10%), and variation in nanoparticle placement (~5%). At volume fractions less than 20% Au, the optical response is insensitive to particle size, defects, and inhomogeneity in the superlattice. At elevated volume fractions (20% and 25%), structures incorporating different sized nanoparticles (10-, 20-, and 40-nm diameter)more » each exhibit distinct far-field extinction and near-field properties. These optical properties are most pronounced in lattices with larger particles, which at fixed volume fraction have greater plasmonic coupling than those with smaller particles. Moreover, the incorporation of experimentally informed inhomogeneity leads to variation in far-field extinction and inconsistent electric-field intensities throughout the lattice, demonstrating that volume fraction is not sufficient to describe the optical properties of such structures. Finally, these data have important implications for understanding the role of particle and lattice inhomogeneity in determining the properties of plasmonic nanoparticle lattices with deliberately designed optical properties.« less

A single molecule study of G-quadruplex and short duplex DNA structures

NASA Astrophysics Data System (ADS)

Roy, William A., Jr.

Given that certain conditions are met, a single stranded DNA/RNA (ssDNA/RNA) structure called G-quadruplex (GQ) can form in regions throughout the genome, including at the telomeres and internal regions of the chromosomes. These structures serve various functions depending on the region in which they form which include protecting the chromosome ends, interfering with telomere elongation in cancer cells, and regulating transcription and translation level gene expression. Due to their high stability, various cellular mechanisms, such as GQ destabilizing proteins, are employed to unfold these structures during DNA replication or repair. Yet, their distinct layered structure has made GQs an attractive drug target in cancer treatment as GQ stabilizing molecules could inhibit telomerase dependent telomere elongation, a mechanism occurring in the majority of cancer cells to avoid senescence and apoptosis. However, proteins or small molecules interact with GQ that is under the influence of various cellular tension mechanisms, including the tension applied by other nearby molecules or the tension due to DNA structure within the chromatin context. Therefore, it is important to characterize the stability of various GQs and their response to interacting molecules when subjected to a tensile force. We employed a novel DNA-based nano tension generator that utilizes the elastic properties of circularized short double-stranded DNA (dsDNA) oligonucleotides to apply tension on the GQ. Since this is a completely new approach, the majority of this thesis was dedicated to proof-of-principle studies that demonstrated the feasibility and functionality of the method.

Spatial confinement induces hairpins in nicked circular DNA

PubMed Central

Japaridze, Aleksandre; Orlandini, Enzo; Smith, Kathleen Beth; Gmür, Lucas; Valle, Francesco; Micheletti, Cristian

2017-01-01

Abstract In living cells, DNA is highly confined in space with the help of condensing agents, DNA binding proteins and high levels of supercoiling. Due to challenges associated with experimentally studying DNA under confinement, little is known about the impact of spatial confinement on the local structure of the DNA. Here, we have used well characterized slits of different sizes to collect high resolution atomic force microscopy images of confined circular DNA with the aim of assessing the impact of the spatial confinement on global and local conformational properties of DNA. Our findings, supported by numerical simulations, indicate that confinement imposes a large mechanical stress on the DNA as evidenced by a pronounced anisotropy and tangent–tangent correlation function with respect to non-constrained DNA. For the strongest confinement we observed nanometer sized hairpins and interwound structures associated with the nicked sites in the DNA sequence. Based on these findings, we propose that spatial DNA confinement in vivo can promote the formation of localized defects at mechanically weak sites that could be co-opted for biological regulatory functions. PMID:28201616

Recent Insight into the Kinetic Mechanisms and Conformational Dynamics of Y-Family DNA Polymerases

PubMed Central

2015-01-01

The kinetic mechanisms by which DNA polymerases catalyze DNA replication and repair have long been areas of active research. Recently discovered Y-family DNA polymerases catalyze the bypass of damaged DNA bases that would otherwise block replicative DNA polymerases and stall replication forks. Unlike DNA polymerases from the five other families, the Y-family DNA polymerases have flexible, solvent-accessible active sites that are able to tolerate various types of damaged template bases and allow for efficient lesion bypass. Their promiscuous active sites, however, also lead to fidelities that are much lower than those observed for other DNA polymerases and give rise to interesting mechanistic properties. Additionally, the Y-family DNA polymerases have several other unique structural features and undergo a set of conformational changes during substrate binding and catalysis different from those observed for replicative DNA polymerases. In recent years, pre-steady-state kinetic methods have been extensively employed to reveal a wealth of information about the catalytic properties of these fascinating noncanonical DNA polymerases. Here, we review many of the recent findings on the kinetic mechanisms of DNA polymerization with undamaged and damaged DNA substrates by the Y-family DNA polymerases, and the conformational dynamics employed by these error-prone enzymes during catalysis. PMID:24716482

Doped Graphene for DNA Analysis: the Electrochemical Signal is Strongly Influenced by the Kind of Dopant and the Nucleobase Structure

PubMed Central

Tian, Huidi; Wang, Lu; Sofer, Zdenek; Pumera, Martin; Bonanni, Alessandra

2016-01-01

Doping graphene with heteroatoms can alter the electronic and electrochemical properties of the starting material. Contrasting properties should be expected when the doping is carried out with electron donating species (n-type dopants) or with electron withdrawing species (p-type dopants). This in turn can have a profound influence on the electroanalytical performance of the doped material being used for the detection of specific probes. Here we investigate the electrochemical oxidation of DNA bases adenine, guanine, thymine and cytosine on two heteroatom-doped graphene platforms namely boron-doped graphene (p-type dopant) and nitrogen-doped graphene (n-type dopant). We found that overall, boron–doped graphene provided the best response in terms of electrochemical signal sensitivity for all bases. This is due to the electron deficiency of boron-doped graphene, which can promote the oxidation of DNA bases, as opposed to nitrogen-doped graphene which possesses an excess of electrons. Moreover, also the structure of the nucleobase was found to have significant influence on the obtained signal. Our study may open new frontiers in the electrochemical detection of DNA bases which is the first step for label-free DNA analysis. PMID:27623951

DNA nanostructure-directed assembly of metal nanoparticle superlattices

NASA Astrophysics Data System (ADS)

Julin, Sofia; Nummelin, Sami; Kostiainen, Mauri A.; Linko, Veikko

2018-05-01

Structural DNA nanotechnology provides unique, well-controlled, versatile, and highly addressable motifs and templates for assembling materials at the nanoscale. These methods to build from the bottom-up using DNA as a construction material are based on programmable and fully predictable Watson-Crick base pairing. Researchers have adopted these techniques to an increasing extent for creating numerous DNA nanostructures for a variety of uses ranging from nanoelectronics to drug-delivery applications. Recently, an increasing effort has been put into attaching nanoparticles (the size range of 1-20 nm) to the accurate DNA motifs and into creating metallic nanostructures (typically 20-100 nm) using designer DNA nanoshapes as molds or stencils. By combining nanoparticles with the superior addressability of DNA-based scaffolds, it is possible to form well-ordered materials with intriguing and completely new optical, plasmonic, electronic, and magnetic properties. This focused review discusses the DNA structure-directed nanoparticle assemblies covering the wide range of different one-, two-, and three-dimensional systems.

Utilizing Molecular Dynamics ' Multipotent Methodologies to Measure Microscopic Motions of DNA Molecules: A Magniloquent Manuscript On DNA's Means and Mannerisms

NASA Astrophysics Data System (ADS)

Kingsland, Addie

DNA is an amazing molecule which is the basic template for all genetics. It is the primary molecule for storing biological information, and has many applications in nanotechnology. Double-stranded DNA may contain mismatched base pairs beyond the Watson-Crick pairs guanine-cytosine and adenine-thymine. To date, no one has found a physical property of base pair mismatches which describes the behavior of naturally occurring mismatch repair enzymes. Many materials properties of DNA are also unknown, for instance, when pulling DNA in different configurations, different energy differences are observed with no obvious reason why. DNA mismatches also affect their local environment, for instance changing the quantum yield of nearby azobenzene moieties. We utilize molecular dynamics computer simulations to study the structure and dynamics for both matched and mismatched base pairs, within both biological and materials contexts, and in both equilibrium and biased dynamics. We show that mismatched pairs shift further in the plane normal to the DNA strand and are more likely to exhibit non-canonical structures, including the e-motif. Base pair mismatches alter their local environment, affecting the trans- to cis- photoisomerization quantum yield of azobenzene, as well as increasing the likelihood of observing the e-motif. We also show that by using simulated data, we can give new insights on theoretical models to calculate the energetics of pulling DNA strands apart. These results, all relatively inexpensive on modern computer hardware, can help guide the design of DNA-based nanotechnologies, as well as give new insights into the functioning of mismatch repair systems in cancer prevention.

Molecular mechanisms by which oxidative DNA damage promotes telomerase activity.

PubMed

Lee, Hui-Ting; Bose, Arindam; Lee, Chun-Ying; Opresko, Patricia L; Myong, Sua

2017-11-16

Telomeres are highly susceptible to oxidative DNA damage, which if left unrepaired can lead to dysregulation of telomere length homeostasis. Here we employed single molecule FRET, single molecule pull-down and biochemical analysis to investigate how the most common oxidative DNA lesions, 8-oxoguanine (8oxoG) and thymine glycol (Tg), regulate the structural properties of telomeric DNA and telomerase extension activity. In contrast to 8oxoG which disrupts the telomeric DNA structure, Tg exhibits substantially reduced perturbation of G-quadruplex folding. As a result, 8oxoG induces high accessibility, whereas Tg retains limited accessibility, of telomeric G-quadruplex DNA to complementary single stranded DNA and to telomere binding protein POT1. Surprisingly, the Tg lesion stimulates telomerase loading and activity to a similar degree as an 8oxoG lesion. We demonstrate that this unexpected stimulation arises from Tg-induced conformational alterations and dynamics in telomeric DNA. Despite impacting structure by different mechanisms, both 8oxoG and Tg enhance telomerase binding and extension activity to the same degree, potentially contributing to oncogenesis. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Insights into the nature of DNA binding of AbrB-like transcription factors

PubMed Central

Sullivan, Daniel M.; Bobay, Benjamin G.; Kojetin, Douglas J.; Thompson, Richele J.; Rance, Mark; Strauch, Mark A.; Cavanagh, John

2008-01-01

Summary Understanding the DNA recognition and binding by the AbrB-like family of transcriptional regulators is of significant interest since these proteins enable bacteria to elicit the appropriate response to diverse environmental stimuli. Although these ‘transition-state regulator’ proteins have been well characterized at the genetic level, the general and specific mechanisms of DNA binding remain elusive. We present RDC-refined NMR solution structures and dynamic properties of the DNA-binding domains of three Bacillus subtilis transition-state regulators AbrB, Abh, and SpoVT. We combined previously investigated DNase I footprinting, DNA methylation, gel shift assays, mutagenic and NMR studies to generate a structural model of the complex between AbrBN55 and its cognate promoter, abrB8. These investigations have enabled us to generate the first model for the specific nature of the transition-state regulator-DNA interaction. PMID:19000822

Structural and thermodynamic principles of viral packaging.

PubMed

Petrov, Anton S; Harvey, Stephen C

2007-01-01

Packaging of genetic material inside a capsid is one of the major processes in the lifecycle of bacteriophages. To establish the basic principles of packing double-stranded DNA into a phage, we present a low-resolution model of bacteriophage varphi29 and report simulations of DNA packaging. The simulations show excellent agreement with available experimental data, including the forces of packaging and the average structures seen in cryo-electron microscopy. The conformation of DNA inside the bacteriophage is primarily determined by the shape of the capsid and the elastic properties of DNA, but the energetics of packaging are dominated by electrostatic repulsions and the large entropic penalty associated with DNA confinement. In this slightly elongated capsid, the DNA assumes a folded toroidal conformation, rather than a coaxial spool. The model can be used to study packaging of other bacteriophages with different shapes under a range of environmental conditions.

The Purine Bias of Coding Sequences is Determined by Physicochemical Constraints on Proteins.

PubMed

Ponce de Leon, Miguel; de Miranda, Antonio Basilio; Alvarez-Valin, Fernando; Carels, Nicolas

2014-01-01

For this report, we analyzed protein secondary structures in relation to the statistics of three nucleotide codon positions. The purpose of this investigation was to find which properties of the ribosome, tRNA or protein level, could explain the purine bias (Rrr) as it is observed in coding DNA. We found that the Rrr pattern is the consequence of a regularity (the codon structure) resulting from physicochemical constraints on proteins and thermodynamic constraints on ribosomal machinery. The physicochemical constraints on proteins mainly come from the hydropathy and molecular weight (MW) of secondary structures as well as the energy cost of amino acid synthesis. These constraints appear through a network of statistical correlations, such as (i) the cost of amino acid synthesis, which is in favor of a higher level of guanine in the first codon position, (ii) the constructive contribution of hydropathy alternation in proteins, (iii) the spatial organization of secondary structure in proteins according to solvent accessibility, (iv) the spatial organization of secondary structure according to amino acid hydropathy, (v) the statistical correlation of MW with protein secondary structures and their overall hydropathy, (vi) the statistical correlation of thymine in the second codon position with hydropathy and the energy cost of amino acid synthesis, and (vii) the statistical correlation of adenine in the second codon position with amino acid complexity and the MW of secondary protein structures. Amino acid physicochemical properties and functional constraints on proteins constitute a code that is translated into a purine bias within the coding DNA via tRNAs. In that sense, the Rrr pattern within coding DNA is the effect of information transfer on nucleotide composition from protein to DNA by selection according to the codon positions. Thus, coding DNA structure and ribosomal machinery co-evolved to minimize the energy cost of protein coding given the functional constraints on proteins.

Nematic DNA Thermotropic Liquid Crystals with Photoresponsive Mechanical Properties.

PubMed

Zhang, Lei; Maity, Sourav; Liu, Kai; Liu, Qing; Göstl, Robert; Portale, Giuseppe; Roos, Wouter H; Herrmann, Andreas

2017-09-01

Over the last decades, water-based lyotropic liquid crystals of nucleic acids have been extensively investigated because of their important role in biology. Alongside, solvent-free thermotropic liquid crystals (TLCs) from DNA are gaining great interest, owing to their relevance to DNA-inspired optoelectronic applications. Up to now, however, only the smectic phase of DNA TLCs has been reported. The development of new mesophases including nematic, hexagonal, and cubic structures for DNA TLCs remains a significant challenge, which thus limits their technological applications considerably. In this work, a new type of DNA TLC that is formed by electrostatic complexation of anionic oligonucleotides and cationic surfactants containing an azobenzene (AZO) moiety is demonstrated. DNA-AZO complexes form a stable nematic mesophase over a temperature range from -7 to 110 °C and retain double-stranded DNA structure at ambient temperature. Photoisomerization of the AZO moieties from the E- to the Z-form alters the stiffness of the DNA-AZO hybrid materials opening a pathway toward the development of DNA TLCs as stimuli-responsive biomaterials. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

DNA Glycosylases Search for and Remove Oxidized DNA Bases

PubMed Central

Wallace, Susan S.

2014-01-01

The following mini review summarizes recent research from the Author’s laboratory as presented to the Environmental Mutagen Society in October 2012. It provides an overview of the DNA glycosylases that recognize oxidized DNA bases using the Fpg/Nei family of DNA glycosylases as models for how structure can inform function. For example, even though human NEIL1 and the plant and fungal orthologs lack the zinc finger shown to be required for binding, DNA crystal structures revealed a “zincless finger” with the same properties. Also the “lesion recognition loop” is not involved in lesion recognition rather stabilization of 8-oxoG in the active site pocket. Unlike the other Fpg/Nei family members, Neil3 lacks two of the three void-filling residues that stabilize the duplex and interact with the opposite strand which may account for its preference for lesions in single stranded DNA. We also showed, using single molecule approaches, that DNA glycosylases search for their substrates in a sea of undamaged DNA by using a wedge residue that is inserted into the DNA helix to probe for the presence of damage. PMID:24123395

Volume and porosity thermal regulation in lipid mesophases by coupling mobile ligands to soft membranes

NASA Astrophysics Data System (ADS)

Parolini, Lucia; Mognetti, Bortolo M.; Kotar, Jurij; Eiser, Erika; Cicuta, Pietro; di Michele, Lorenzo

2015-01-01

Short DNA linkers are increasingly being exploited for driving-specific self-assembly of Brownian objects. DNA-functionalized colloids can assemble into ordered or amorphous materials with tailored morphology. Recently, the same approach has been applied to compliant units, including emulsion droplets and lipid vesicles. The liquid structure of these substrates introduces new degrees of freedom: the tethers can diffuse and rearrange, radically changing the physics of the interactions. Unlike droplets, vesicles are extremely deformable and DNA-mediated adhesion causes significant shape adjustments. We investigate experimentally the thermal response of pairs and networks of DNA-tethered liposomes and observe two intriguing and possibly useful collective properties: negative thermal expansion and tuneable porosity of the liposome networks. A model providing a thorough understanding of this unexpected phenomenon is developed, explaining the emergent properties out of the interplay between the temperature-dependent deformability of the vesicles and the DNA-mediated adhesive forces.

Volume and porosity thermal regulation in lipid mesophases by coupling mobile ligands to soft membranes

PubMed Central

Parolini, Lucia; Mognetti, Bortolo M.; Kotar, Jurij; Eiser, Erika; Cicuta, Pietro; Di Michele, Lorenzo

2015-01-01

Short DNA linkers are increasingly being exploited for driving-specific self-assembly of Brownian objects. DNA-functionalized colloids can assemble into ordered or amorphous materials with tailored morphology. Recently, the same approach has been applied to compliant units, including emulsion droplets and lipid vesicles. The liquid structure of these substrates introduces new degrees of freedom: the tethers can diffuse and rearrange, radically changing the physics of the interactions. Unlike droplets, vesicles are extremely deformable and DNA-mediated adhesion causes significant shape adjustments. We investigate experimentally the thermal response of pairs and networks of DNA-tethered liposomes and observe two intriguing and possibly useful collective properties: negative thermal expansion and tuneable porosity of the liposome networks. A model providing a thorough understanding of this unexpected phenomenon is developed, explaining the emergent properties out of the interplay between the temperature-dependent deformability of the vesicles and the DNA-mediated adhesive forces. PMID:25565580

DNA-linked NanoParticle Lattices with Diamond Symmetry: Stability, Shape and Optical Properties

NASA Astrophysics Data System (ADS)

Emamy, Hamed; Tkachenko, Alexei; Gang, Oleg; Starr, Francis

The linking of nanoparticles (NP) by DNA has been proven to be an effective means to create NP lattices with specific order. Lattices with diamond symmetry are predicted to offer novel photonic properties, but self-assembly of such lattices has proven to be challenging due to the low packing fraction, sensitivity to bond orientation, and local heterogeneity. Recently, we reported an approach to create diamond NP lattices based on the association between anisotropic particles with well-defined tetravalent DNA binding topology and isotropically functionalized NP. Here, we use molecular dynamics simulations to evaluate the Gibbs free energy of these lattices, and thereby determine the stability of these lattices as a function of NP size and DNA stiffness. We also predict the equilibrium shape for the cubic diamond crystallite using the Wulff construction method. Specifically, we predict the equilibrium shape using the surface energy for different crystallographic planes. We evaluate surface energy directly form molecular dynamics simulation, which we correlate with theoretical estimates from the expected number of broken DNA bonds along a facet. Furthermore we study the optical properties of this structure, e.g optical bandgap.

Mechanical properties of DNA-like polymers

PubMed Central

Peters, Justin P.; Yelgaonkar, Shweta P.; Srivatsan, Seergazhi G.; Tor, Yitzhak; James Maher, L.

2013-01-01

The molecular structure of the DNA double helix has been known for 60 years, but we remain surprisingly ignorant of the balance of forces that determine its mechanical properties. The DNA double helix is among the stiffest of all biopolymers, but neither theory nor experiment has provided a coherent understanding of the relative roles of attractive base stacking forces and repulsive electrostatic forces creating this stiffness. To gain insight, we have created a family of double-helical DNA-like polymers where one of the four normal bases is replaced with various cationic, anionic or neutral analogs. We apply DNA ligase-catalyzed cyclization kinetics experiments to measure the bending and twisting flexibilities of these polymers under low salt conditions. Interestingly, we show that these modifications alter DNA bending stiffness by only 20%, but have much stronger (5-fold) effects on twist flexibility. We suggest that rather than modifying DNA stiffness through a mechanism easily interpretable as electrostatic, the more dominant effect of neutral and charged base modifications is their ability to drive transitions to helical conformations different from canonical B-form DNA. PMID:24013560

A Three-State System Based on Branched DNA Hybrids.

PubMed

He, Shiliang; Richert, Clemens

2018-03-26

There is a need for materials that respond to chemical or physical stimuli through a change in their structure. While a transition between water-soluble form and solid is not uncommon for DNA-based structures, systems that transition between three different states at room temperature and ambient pressure are rare. Here we report the preparation of branched DNA hybrids with eight oligodeoxycytidylate arms via solution-phase, H-phosphonate-based synthesis. Some hybrids assemble into hydrogels upon lowering the pH, acting as efficient gelators at pH 4-6, but can also transition into a more condensed solid state form upon exposure to divalent cations. Together with the homogeneous solutions that the i-motif-forming compounds give at neutral pH, three-state systems result. Each state has its own color, if chromophores are included in the system. The assembly and gelation properties can be tuned by choosing the chain length of the arms. Their responsive properties make the dC-rich DNA hybrids candidates for smart material applications. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Structural properties of oligonucleotide monolayers on gold surfaces probed by fluorescence investigations.

PubMed

Rant, Ulrich; Arinaga, Kenji; Fujita, Shozo; Yokoyama, Naoki; Abstreiter, Gerhard; Tornow, Marc

2004-11-09

We present optical investigations on the conformation of oligonucleotide layers on Au surfaces. Our studies concentrate on the effect of varying surface coverage densities on the structural properties of layers of 12- and 24mer single-stranded DNA, tethered to the Au surface at one end while being labeled with a fluorescent marker at the opposing end. The distance-dependent energy transfer from the marker dye to the metal surface, which causes quenching of the observed fluorescence, is used to provide information on the orientation of the DNA strands relative to the surface. Variations in the oligonucleotide coverage density, as determined from electrochemical quantification, over 2 orders of magnitude are achieved by employing different preparation conditions. The observed enhancement in fluorescence intensity with increasing DNA coverage can be related to a model involving mutual steric interactions of oligonucleotides on the surface, as well as fluorescence quenching theory. Finally, the applicability of the presented concepts for investigations of heterogeneous monolayers is demonstrated by means of studying the coadsorption of mercaptohexanol onto DNA-modified Au surfaces.

Molecular self-assembly using peptide nucleic acids.

PubMed

Berger, Or; Gazit, Ehud

2017-01-01

Peptide nucleic acids (PNAs) are extensively studied for the control of genetic expression since their design in the 1990s. However, the application of PNAs in nanotechnology is much more recent. PNAs share the specific base-pair recognition characteristic of DNA together with material-like properties of polyamides, both proteins and synthetic polymers, such as Kevlar and Nylon. The first application of PNA was in the form of PNA-amphiphiles, resulting in the formation of either lipid integrated structures, hydrogels or fibrillary assemblies. Heteroduplex DNA-PNA assemblies allow the formation of hybrid structures with higher stability as compared with pure DNA. A systematic screen for minimal PNA building blocks resulted in the identification of guanine-containing di-PNA assemblies and protected guanine-PNA monomer spheres showing unique optical properties. Finally, the co-assembly of PNA with thymine-like three-faced cyanuric acid allowed the assembly of poly-adenine PNA into fibers. In summary, we believe that PNAs represent a new and important family of building blocks which converges the advantages of both DNA- and peptide-nanotechnologies. © 2016 Wiley Periodicals, Inc.

An anti-DNA antibody prefers damaged dsDNA over native.

PubMed

Akberova, N I; Zhmurov, A A; Nevzorova, T A; Litvinov, R I

2017-01-01

DNA-protein interactions, including DNA-antibody complexes, have both fundamental and practical significance. In particular, antibodies against double-stranded DNA play an important role in the pathogenesis of autoimmune diseases. Elucidation of structural mechanisms of an antigen recognition and interaction of anti-DNA antibodies provides a basis for understanding the role of DNA-containing immune complexes in human pathologies and for new treatments. Here we used Molecular Dynamic simulations of bimolecular complexes of a segment of dsDNA with a monoclonal anti-DNA antibody's Fab-fragment to obtain detailed structural and physical characteristics of the dynamic intermolecular interactions. Using a computationally modified crystal structure of a Fab-DNA complex (PDB: 3VW3), we studied in silico equilibrium Molecular Dynamics of the Fab-fragment associated with two homologous dsDNA fragments, containing or not containing dimerized thymine, a product of DNA photodamage. The Fab-fragment interactions with the thymine dimer-containing DNA was thermodynamically more stable than with the native DNA. The amino acid residues constituting a paratope and the complementary nucleotide epitopes for both Fab-DNA constructs were identified. Stacking and electrostatic interactions were shown to play the main role in the antibody-dsDNA contacts, while hydrogen bonds were less significant. The aggregate of data show that the chemically modified dsDNA (containing a covalent thymine dimer) has a higher affinity toward the antibody and forms a stronger immune complex. These findings provide a mechanistic insight into formation and properties of the pathogenic anti-DNA antibodies in autoimmune diseases, such as systemic lupus erythematosus, associated with skin photosensibilization and DNA photodamage.

6-Thioguanine alters the structure and stability of duplex DNA and inhibits quadruplex DNA formation.

PubMed

Marathias, V M; Sawicki, M J; Bolton, P H

1999-07-15

The ability to chemically synthesize biomolecules has opened up the opportunity to observe changes in structure and activity that occur upon single atom substitution. In favorable cases this can provide information about the roles of individual atoms. The substitution of 6-thioguanine (6SG) for guanine is a potentially very useful single atom substitution as 6SG has optical, photocrosslinking, metal ion binding and other properties of potential utility. In addition, 6-mercaptopurine is a clinically important pro-drug that is activated by conversion into 6SG by cells. The results presented here indicate that the presence of 6SG blocks the formation of quadruplex DNA. The presence of 6SG alters the structure and lowers the thermal stability of duplex DNA, but duplex DNA can be formed in the presence of 6SG. These results indicate that some of the cytotoxic activity of 6SG may be due to disruption of the quadruplex structures formed by telomere and other DNAs. This additional mode of action is consistent with the delayed onset of cytotoxicity.

A base-modified PNA-graphene oxide platform as a turn-on fluorescence sensor for the detection of human telomeric repeats

NASA Astrophysics Data System (ADS)

Sabale, Pramod M.; George, Jerrin Thomas; Srivatsan, Seergazhi G.

2014-08-01

Given the biological and therapeutic significance of telomeres and other G-quadruplex forming sequences in human genome, it is highly desirable to develop simple methods to study these structures, which can also be implemented in screening formats for the discovery of G-quadruplex binders. The majority of telomere detection methods developed so far are laborious and use elaborate assay and instrumental setups, and hence, are not amenable to discovery platforms. Here, we describe the development of a simple homogeneous fluorescence turn-on method, which uses a unique combination of an environment-sensitive fluorescent nucleobase analogue, the superior base pairing property of PNA, and DNA-binding and fluorescence quenching properties of graphene oxide, to detect human telomeric DNA repeats of varying lengths. Our results demonstrate that this method, which does not involve a rigorous assay setup, would provide new opportunities to study G-quadruplex structures.Given the biological and therapeutic significance of telomeres and other G-quadruplex forming sequences in human genome, it is highly desirable to develop simple methods to study these structures, which can also be implemented in screening formats for the discovery of G-quadruplex binders. The majority of telomere detection methods developed so far are laborious and use elaborate assay and instrumental setups, and hence, are not amenable to discovery platforms. Here, we describe the development of a simple homogeneous fluorescence turn-on method, which uses a unique combination of an environment-sensitive fluorescent nucleobase analogue, the superior base pairing property of PNA, and DNA-binding and fluorescence quenching properties of graphene oxide, to detect human telomeric DNA repeats of varying lengths. Our results demonstrate that this method, which does not involve a rigorous assay setup, would provide new opportunities to study G-quadruplex structures. Electronic supplementary information (ESI) available. Figures, tables, experimental procedures and NMR spectra. See DOI: 10.1039/c4nr00878b

Predicting and analyzing DNA-binding domains using a systematic approach to identifying a set of informative physicochemical and biochemical properties

PubMed Central

2011-01-01

Background Existing methods of predicting DNA-binding proteins used valuable features of physicochemical properties to design support vector machine (SVM) based classifiers. Generally, selection of physicochemical properties and determination of their corresponding feature vectors rely mainly on known properties of binding mechanism and experience of designers. However, there exists a troublesome problem for designers that some different physicochemical properties have similar vectors of representing 20 amino acids and some closely related physicochemical properties have dissimilar vectors. Results This study proposes a systematic approach (named Auto-IDPCPs) to automatically identify a set of physicochemical and biochemical properties in the AAindex database to design SVM-based classifiers for predicting and analyzing DNA-binding domains/proteins. Auto-IDPCPs consists of 1) clustering 531 amino acid indices in AAindex into 20 clusters using a fuzzy c-means algorithm, 2) utilizing an efficient genetic algorithm based optimization method IBCGA to select an informative feature set of size m to represent sequences, and 3) analyzing the selected features to identify related physicochemical properties which may affect the binding mechanism of DNA-binding domains/proteins. The proposed Auto-IDPCPs identified m=22 features of properties belonging to five clusters for predicting DNA-binding domains with a five-fold cross-validation accuracy of 87.12%, which is promising compared with the accuracy of 86.62% of the existing method PSSM-400. For predicting DNA-binding sequences, the accuracy of 75.50% was obtained using m=28 features, where PSSM-400 has an accuracy of 74.22%. Auto-IDPCPs and PSSM-400 have accuracies of 80.73% and 82.81%, respectively, applied to an independent test data set of DNA-binding domains. Some typical physicochemical properties discovered are hydrophobicity, secondary structure, charge, solvent accessibility, polarity, flexibility, normalized Van Der Waals volume, pK (pK-C, pK-N, pK-COOH and pK-a(RCOOH)), etc. Conclusions The proposed approach Auto-IDPCPs would help designers to investigate informative physicochemical and biochemical properties by considering both prediction accuracy and analysis of binding mechanism simultaneously. The approach Auto-IDPCPs can be also applicable to predict and analyze other protein functions from sequences. PMID:21342579

Characterization of Nanofluidic Entropic Trap Array for DNA Separation

NASA Astrophysics Data System (ADS)

Han, Jongyoon

2003-03-01

Micromachined nanoscale fluidic structures can provide new opportunities in biomolecule manipulation and sorting, because their chemical and physical properties can be controlled easily unlike random nanoporous materials. As an example of regular nanostructures used for biomolecule manipulation and sorting, a nanofluidic entropic trap array for DNA separation is presented. Nanofluidic channels as thin as 75nm were used as a molecular sieve instead of agarose gel for DNA separation. The interaction between DNA molecules and the nanofluidic structure determines the DNA migration speed, which was used to separate DNA molecules in a dc electrophoresis. Separation of long DNA (up to 200kbp) has been achieved within 30 minutes, using less than a picogram quantities of DNA, with only 1.5cm long channels.[1] In addition to the efficiency improvement, nanofluidic DNA entropic traps have a regular structure that can be easily modeled theoretically. The theoretical model could be the basis for improving the system performance for further optimization in separation size range and resolution. The process of DNA moving out of the entropic trap was theoretically modeled, and the prediction of the theoretical model was compared with the experimental data.[2] The selectivity, resolution, and the separation range of DNA for a given entropic trap separation system was discussed in terms of the number of entropic traps, various structural parameters of the system, and the electric field. It is expected that this system could be used for analyzing a small amount of ultra-long DNA molecules. (1) Han, J.; Craighead, H. G. Science 2000, 288, 1026-1029. (2) Han, J.; Craighead, H. G. Anal. Chem. 2002, 74, 394-401.

An open reading frame in intron seven of the sea urchin DNA-methyltransferase gene codes for a functional AP1 endonuclease.

PubMed

Cioffi, Anna Valentina; Ferrara, Diana; Cubellis, Maria Vittoria; Aniello, Francesco; Corrado, Marcella; Liguori, Francesca; Amoroso, Alessandro; Fucci, Laura; Branno, Margherita

2002-08-01

Analysis of the genome structure of the Paracentrotus lividus (sea urchin) DNA methyltransferase (DNA MTase) gene showed the presence of an open reading frame, named METEX, in intron 7 of the gene. METEX expression is developmentally regulated, showing no correlation with DNA MTase expression. In fact, DNA MTase transcripts are present at high concentrations in the early developmental stages, while METEX is expressed at late stages of development. Two METEX cDNA clones (Met1 and Met2) that are different in the 3' end have been isolated in a cDNA library screening. The putative translated protein from Met2 cDNA clone showed similarity with Escherichia coli endonuclease III on the basis of sequence and predictive three-dimensional structure. The protein, overexpressed in E. coli and purified, had functional properties similar to the endonuclease specific for apurinic/apyrimidinic (AP) sites on the basis of the lyase activity. Therefore the open reading frame, present in intron 7 of the P. lividus DNA MTase gene, codes for a functional AP endonuclease designated SuAP1.

Structural and Functional Studies of H. seropedicae RecA Protein - Insights into the Polymerization of RecA Protein as Nucleoprotein Filament.

PubMed

Leite, Wellington C; Galvão, Carolina W; Saab, Sérgio C; Iulek, Jorge; Etto, Rafael M; Steffens, Maria B R; Chitteni-Pattu, Sindhu; Stanage, Tyler; Keck, James L; Cox, Michael M

2016-01-01

The bacterial RecA protein plays a role in the complex system of DNA damage repair. Here, we report the functional and structural characterization of the Herbaspirillum seropedicae RecA protein (HsRecA). HsRecA protein is more efficient at displacing SSB protein from ssDNA than Escherichia coli RecA protein. HsRecA also promotes DNA strand exchange more efficiently. The three dimensional structure of HsRecA-ADP/ATP complex has been solved to 1.7 Å resolution. HsRecA protein contains a small N-terminal domain, a central core ATPase domain and a large C-terminal domain, that are similar to homologous bacterial RecA proteins. Comparative structural analysis showed that the N-terminal polymerization motif of archaeal and eukaryotic RecA family proteins are also present in bacterial RecAs. Reconstruction of electrostatic potential from the hexameric structure of HsRecA-ADP/ATP revealed a high positive charge along the inner side, where ssDNA is bound inside the filament. The properties of this surface may explain the greater capacity of HsRecA protein to bind ssDNA, forming a contiguous nucleoprotein filament, displace SSB and promote DNA exchange relative to EcRecA. Our functional and structural analyses provide insight into the molecular mechanisms of polymerization of bacterial RecA as a helical nucleoprotein filament.

Irradiation effects on electrical properties of DNA solution/Al Schottky diodes

NASA Astrophysics Data System (ADS)

Al-Ta'ii, Hassan Maktuff Jaber; Periasamy, Vengadesh; Iwamoto, Mitsumasa

2018-04-01

Deoxyribonucleic acid (DNA) has emerged as one of the most exciting organic material and as such extensively studied as a smart electronic material since the last few decades. DNA molecules have been reported to be utilized in the fabrication of small-scaled sensors and devices. In this current work, the effect of alpha radiation on the electrical properties of an Al/DNA/Al device using DNA solution was studied. It was observed that the carrier transport was governed by electrical interface properties at the Al-DNA interface. Current ( I)-voltage ( V) curves were analyzed by employing the interface limited Schottky current equations, i.e., conventional and Cheung and Cheung's models. Schottky parameters such as ideality factor, barrier height and series resistance were also determined. The extracted barrier height of the Schottky contact before and after radiation was calculated as 0.7845, 0.7877, 0.7948 and 0.7874 eV for the non-radiated, 12, 24 and 36 mGy, respectively. Series resistance of the structure was found to decline with the increase in the irradiation, which was due to the increase in the free radical root effects in charge carriers in the DNA solution. Results pertaining to the electronic profiles obtained in this work may provide a better understanding for the development of precise and rapid radiation sensors using DNA solution.

Structural and Functional Basis for an EBNA1 Hexameric Ring in Epstein-Barr Virus Episome Maintenance.

PubMed

Deakyne, Julianna S; Malecka, Kimberly A; Messick, Troy E; Lieberman, Paul M

2017-10-01

Epstein-Barr virus (EBV) establishes a stable latent infection that can persist for the life of the host. EBNA1 is required for the replication, maintenance, and segregation of the latent episome, but the structural features of EBNA1 that confer each of these functions are not completely understood. Here, we have solved the X-ray crystal structure of an EBNA1 DNA-binding domain (DBD) and discovered a novel hexameric ring oligomeric form. The oligomeric interface pivoted around residue T585 as a joint that links and stabilizes higher-order EBNA1 complexes. Substitution mutations around the interface destabilized higher-order complex formation and altered the cooperative DNA-binding properties of EBNA1. Mutations had both positive and negative effects on EBNA1-dependent DNA replication and episome maintenance with OriP. We found that one naturally occurring polymorphism in the oligomer interface (T585P) had greater cooperative DNA binding in vitro , minor defects in DNA replication, and pronounced defects in episome maintenance. The T585P mutant was compromised for binding to OriP in vivo as well as for assembling the origin recognition complex subunit 2 (ORC2) and trimethylated histone 3 lysine 4 (H3K4me3) at OriP. The T585P mutant was also compromised for forming stable subnuclear foci in living cells. These findings reveal a novel oligomeric structure of EBNA1 with an interface subject to naturally occurring polymorphisms that modulate EBNA1 functional properties. We propose that EBNA1 dimers can assemble into higher-order oligomeric structures important for diverse functions of EBNA1. IMPORTANCE Epstein-Barr virus is a human gammaherpesvirus that is causally associated with various cancers. Carcinogenic properties are linked to the ability of the virus to persist in the latent form for the lifetime of the host. EBNA1 is a sequence-specific DNA-binding protein that is consistently expressed in EBV tumors and is the only viral protein required to maintain the viral episome during latency. The structural and biochemical mechanisms by which EBNA1 allows the long-term persistence of the EBV genome are currently unclear. Here, we have solved the crystal structure of an EBNA1 hexameric ring and characterized key residues in the interface required for higher-order complex formation and long-term plasmid maintenance. Copyright © 2017 American Society for Microbiology.

The Incorporation of Ribonucleotides Induces Structural and Conformational Changes in DNA.

PubMed

Meroni, Alice; Mentegari, Elisa; Crespan, Emmanuele; Muzi-Falconi, Marco; Lazzaro, Federico; Podestà, Alessandro

2017-10-03

Ribonucleotide incorporation is the most common error occurring during DNA replication. Cells have hence developed mechanisms to remove ribonucleotides from the genome and restore its integrity. Indeed, the persistence of ribonucleotides into DNA leads to severe consequences, such as genome instability and replication stress. Thus, it becomes important to understand the effects of ribonucleotides incorporation, starting from their impact on DNA structure and conformation. Here we present a systematic study of the effects of ribonucleotide incorporation into DNA molecules. We have developed, to our knowledge, a new method to efficiently synthesize long DNA molecules (hundreds of basepairs) containing ribonucleotides, which is based on a modified protocol for the polymerase chain reaction. By means of atomic force microscopy, we could therefore investigate the changes, upon ribonucleotide incorporation, of the structural and conformational properties of numerous DNA populations at the single-molecule level. Specifically, we characterized the scaling of the contour length with the number of basepairs and the scaling of the end-to-end distance with the curvilinear distance, the bending angle distribution, and the persistence length. Our results revealed that ribonucleotides affect DNA structure and conformation on scales that go well beyond the typical dimension of the single ribonucleotide. In particular, the presence of ribonucleotides induces a systematic shortening of the molecules, together with a decrease of the persistence length. Such structural changes are also likely to occur in vivo, where they could directly affect the downstream DNA transactions, as well as interfere with protein binding and recognition. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Structural properties of prokaryotic promoter regions correlate with functional features.

PubMed

Meysman, Pieter; Collado-Vides, Julio; Morett, Enrique; Viola, Roberto; Engelen, Kristof; Laukens, Kris

2014-01-01

The structural properties of the DNA molecule are known to play a critical role in transcription. In this paper, the structural profiles of promoter regions were studied within the context of their diversity and their function for eleven prokaryotic species; Escherichia coli, Klebsiella pneumoniae, Salmonella Typhimurium, Pseudomonas auroginosa, Geobacter sulfurreducens Helicobacter pylori, Chlamydophila pneumoniae, Synechocystis sp., Synechoccocus elongates, Bacillus anthracis, and the archaea Sulfolobus solfataricus. The main anchor point for these promoter regions were transcription start sites identified through high-throughput experiments or collected within large curated databases. Prokaryotic promoter regions were found to be less stable and less flexible than the genomic mean across all studied species. However, direct comparison between species revealed differences in their structural profiles that can not solely be explained by the difference in genomic GC content. In addition, comparison with functional data revealed that there are patterns in the promoter structural profiles that can be linked to specific functional loci, such as sigma factor regulation or transcription factor binding. Interestingly, a novel structural element clearly visible near the transcription start site was found in genes associated with essential cellular functions and growth in several species. Our analyses reveals the great diversity in promoter structural profiles both between and within prokaryotic species. We observed relationships between structural diversity and functional features that are interesting prospects for further research to yet uncharacterized functional loci defined by DNA structural properties.

Differential Deformability of the DNA Minor Groove and Altered BI/BII Backbone Conformational Equilibrium by the Monovalent Ions Li+, Na+, K+ and Rb+ via Water-Mediated Hydrogen Bonding

PubMed Central

Savelyev, Alexey; MacKerell, Alexander D.

2015-01-01

Recently, we reported the differential impact of the monovalent cations Li+, Na+, K+ and Rb+ on DNA conformational properties. These were identified from variations in the calculated solution-state X-ray DNA spectra as a function of the ion type in the solvation buffer in MD simulations using our recently developed polarizable force field based on the classical Drude oscillator. Changes in the DNA structure were found to mainly involve variations in the minor groove width. Because minor groove dimensions vary significantly in protein-DNA complexes and have been shown to play a critical role in both specific and nonspecific DNA readout, understanding the origins of the observed differential DNA modulation by the first-group monovalent ions is of great biological importance. In the present study we show that the primary microscopic mechanism for the phenomenon is the formation of the water-mediated hydrogen bonds between solvated cations located inside the minor groove and simultaneously to two DNA strands, a process whose intensity and impact on DNA structure depends on both the type of the ion and DNA sequence. Additionally, it is shown that formation of such ion-DNA hydrogen bond complexes appreciably modulates the conformation of the backbone by increasing the population of the BII substate. Notably, the differential impact of the ions on DNA conformational behavior is only predicted by the Drude polarizable model for DNA, with virtually no effect observed from MD simulations utilizing the additive CHARMM36 model. Analysis of dipole moments of the water shows the Drude SWM4 model to possess high sensitivity to changes in the local environment, which indicates the important role of electronic polarization in the salt-dependent conformational properties. This also suggests that inclusion of polarization effects is required to model even relatively simple biological systems such as DNA in various ionic solutions. PMID:26575937

Triplex-forming oligonucleotides: a third strand for DNA nanotechnology

PubMed Central

2018-01-01

Abstract DNA self-assembly has proved to be a useful bottom-up strategy for the construction of user-defined nanoscale objects, lattices and devices. The design of these structures has largely relied on exploiting simple base pairing rules and the formation of double-helical domains as secondary structural elements. However, other helical forms involving specific non-canonical base-base interactions have introduced a novel paradigm into the process of engineering with DNA. The most notable of these is a three-stranded complex generated by the binding of a third strand within the duplex major groove, generating a triple-helical (‘triplex’) structure. The sequence, structural and assembly requirements that differentiate triplexes from their duplex counterparts has allowed the design of nanostructures for both dynamic and/or structural purposes, as well as a means to target non-nucleic acid components to precise locations within a nanostructure scaffold. Here, we review the properties of triplexes that have proved useful in the engineering of DNA nanostructures, with an emphasis on applications that hitherto have not been possible by duplex formation alone. PMID:29228337

Understanding the mechanisms of protein-DNA interactions

NASA Astrophysics Data System (ADS)

Lavery, Richard

2004-03-01

Structural, biochemical and thermodynamic data on protein-DNA interactions show that specific recognition cannot be reduced to a simple set of binary interactions between the partners (such as hydrogen bonds, ion pairs or steric contacts). The mechanical properties of the partners also play a role and, in the case of DNA, variations in both conformation and flexibility as a function of base sequence can be a significant factor in guiding a protein to the correct binding site. All-atom molecular modeling offers a means of analyzing the role of different binding mechanisms within protein-DNA complexes of known structure. This however requires estimating the binding strengths for the full range of sequences with which a given protein can interact. Since this number grows exponentially with the length of the binding site it is necessary to find a method to accelerate the calculations. We have achieved this by using a multi-copy approach (ADAPT) which allows us to build a DNA fragment with a variable base sequence. The results obtained with this method correlate well with experimental consensus binding sequences. They enable us to show that indirect recognition mechanisms involving the sequence dependent properties of DNA play a significant role in many complexes. This approach also offers a means of predicting protein binding sites on the basis of binding energies, which is complementary to conventional lexical techniques.

Investigation of arc repressor DNA-binding specificity by comparative molecular dynamics simulations.

PubMed

Song, Wei; Guo, Jun-Tao

2015-01-01

Transcription factors regulate gene expression through binding to specific DNA sequences. How transcription factors achieve high binding specificity is still not well understood. In this paper, we investigated the role of protein flexibility in protein-DNA-binding specificity by comparative molecular dynamics (MD) simulations. Protein flexibility has been considered as a key factor in molecular recognition, which is intrinsically a dynamic process involving fine structural fitting between binding components. In this study, we performed comparative MD simulations on wild-type and F10V mutant P22 Arc repressor in both free and complex conformations. The F10V mutant has lower DNA-binding specificity though both the bound and unbound main-chain structures between the wild-type and F10V mutant Arc are highly similar. We found that the DNA-binding motif of wild-type Arc is structurally more flexible than the F10V mutant in the unbound state, especially for the six DNA base-contacting residues in each dimer. We demonstrated that the flexible side chains of wild-type Arc lead to a higher DNA-binding specificity through forming more hydrogen bonds with DNA bases upon binding. Our simulations also showed a possible conformational selection mechanism for Arc-DNA binding. These results indicate the important roles of protein flexibility and dynamic properties in protein-DNA-binding specificity.

Mechanical properties of transription

NASA Astrophysics Data System (ADS)

Sevier, Stuart; Levine, Herbert

Over the last several decades it has been increasingly recognized that both stochastic and mechanical processes play a central role in transcription. Though many aspects have been explained a number of fundamental properties are undeveloped. Recent results have pointed to mechanical feedback as the source of transcriptional bursting and DNA supercoiling but a reconciliation of this perspective with preexisting views of transcriptional is lacking. In this work we present a simple model of transcription where RNA elongation, RNA polymerase rotation and DNA supercoiling are coupled. The mechanical properties of each object form a foundational framework for understanding the physical nature of transcription. The resulting model can explain several important aspects of chromatin structure and generates a number of predictions for the mechanical properties of transcription.

Beyond DNA origami: A look on the bright future of nucleic acid nanotechnology

PubMed Central

Michelotti, Nicole; Johnson-Buck, Alexander; Manzo, Anthony J.

2012-01-01

Nucleic acid nanotechnology exploits the programmable molecular recognition properties of natural and synthetic nucleic acids to assemble structures with nanometer-scale precision. In 2006, DNA origami transformed the field by providing a versatile platform for self-assembly of arbitrary shapes from one long DNA strand held in place by hundreds of short, site-specific (spatially addressable) DNA ”staples”. This revolutionary approach has led to the creation of a multitude of 2D and 3D scaffolds that form the basis for functional nanodevices. Not limited to nucleic acids, these nanodevices can incorporate other structural and functional materials, such as proteins and nanoparticles, making them broadly useful for current and future applications in emerging fields such as nanomedicine, nanoelectronics, and alternative energy. PMID:22131292

Precisely Tailored DNA Nanostructures and their Theranostic Applications.

PubMed

Zhu, Bing; Wang, Lihua; Li, Jiang; Fan, Chunhai

2017-12-01

A critical challenge in nanotechnology is the limited precision and controllability of the structural parameters, which brings about concerns in uniformity, reproducibility and performance. Self-assembled DNA nanostructures, as a newly emerged type of nano-biomaterials, possess low-nanometer precision, excellent programmability and addressability. They can precisely arrange various molecules and materials to form spatially ordered complex, resulting in unambiguous physical or chemical properties. Because of these, DNA nanostructures have shown great promise in numerous biomedical theranostic applications. In this account, we briefly review the history and advances on construction of DNA nanoarchitectures and superstructures with accurate structural parameters. We focus on recent progress in exploiting these DNA nanostructures as platforms for quantitative biosensing, intracellular diagnosis, imaging, and smart drug delivery. We also discuss key challenges in practical applications. © 2017 The Chemical Society of Japan & Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Ultrasensitive electrochemical cocaine biosensor based on reversible DNA nanostructure.

PubMed

Sheng, Qinglin; Liu, Ruixiao; Zhang, Sai; Zheng, Jianbin

2014-01-15

We proposed an ultrasensitive electrochemical cocaine biosensor based on the three-dimensional (3D) DNA structure conversion of nanostructure from Triangular Pyramid Frustum (TPFDNA) to Equilateral Triangle (ETDNA). The presence of cocaine triggered the aptamer-composed DNA nanostructure change from "Close" to "Open", leading to obvious faradaic impedance changes. The unique properties with excellent stability and specific rigid structure of the 3D DNA nanostructure made the biosensing functions stable, sensitive, and regenerable. The Faradaic impedance responses were linearly related to cocaine concentration between 1.0 nM and 2.0 μM with a correlation coefficient of 0.993. The limit of detection was calculated to be 0.21 nM following IUPAC recommendations (3Sb/b). It is expected that the distinctive features of DNA nanostructure would make it potentially advantageous for a broad range of biosensing, bionanoelectronics, and therapeutic applications. Copyright © 2013 Elsevier B.V. All rights reserved.

Structural DNA nanotechnology for intelligent drug delivery.

PubMed

Chao, Jie; Liu, Huajie; Su, Shao; Wang, Lianhui; Huang, Wei; Fan, Chunhai

2014-11-01

Drug delivery carriers have been popularly employed to improve solubility, stability, and efficacy of chemical and biomolecular drugs. Despite the rapid progress in this field, it remains a great challenge to develop an ideal carrier with minimal cytotoxicity, high biocompatibility and intelligence for targeted controlled release. The emergence of DNA nanotechnology offers unprecedented opportunities in this regard. Due to the unparalleled self-recognition properties of DNA molecules, it is possible to create numerous artificial DNA nanostructures with well-defined structures and DNA nanodevices with precisely controlled motions. More importantly, recent studies have proven that DNA nanostructures possess greater permeability to the membrane barrier of cells, which pave the way to developing new drug delivery carriers with nucleic acids, are summarized. In this Concept, recent advances on the design and fabrication of both static and dynamic DNA nanostructures, and the use of these nanostructures for the delivery of various types of drugs, are highlighted. It is also demonstrated that dynamic DNA nanostructures provide the required intelligence to realize logically controlled drug release. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Fluorescent DNA-templated silver nanoclusters

NASA Astrophysics Data System (ADS)

Lin, Ruoqian

Because of the ultra-small size and biocompatibility of silver nanoclusters, they have attracted much research interest for their applications in biolabeling. Among the many ways of synthesizing silver nanoclusters, DNA templated method is particularly attractive---the high tunability of DNA sequences provides another degree of freedom for controlling the chemical and photophysical properties. However, systematic studies about how DNA sequences and concentrations are controlling the photophysical properties are still lacking. The aim of this thesis is to investigate the binding mechanisms of silver clusters binding and single stranded DNAs. Here in this thesis, we report synthesis and characterization of DNA-templated silver nanoclusters and provide a systematic interrogation of the effects of DNA concentrations and sequences, including lengths and secondary structures. We performed a series of syntheses utilizing five different sequences to explore the optimal synthesis condition. By characterizing samples with UV-vis and fluorescence spectroscopy, we achieved the most proper reactants ratio and synthesis conditions. Two of them were chosen for further concentration dependence studies and sequence dependence studies. We found that cytosine-rich sequences are more likely to produce silver nanoclusters with stronger fluorescence signals; however, sequences with hairpin secondary structures are more capable in stabilizing silver nanoclusters. In addition, the fluorescence peak emission intensities and wavelengths of the DNA templated silver clusters have sequence dependent fingerprints. This potentially can be applied to sequence sensing in the future. However all the current conclusions are not warranted; there is still difficulty in formulating general rules in DNA strand design and silver nanocluster production. Further investigation of more sequences could solve these questions in the future.

Comprehensive analysis of DNA polymerase III α subunits and their homologs in bacterial genomes

PubMed Central

Timinskas, Kęstutis; Balvočiūtė, Monika; Timinskas, Albertas; Venclovas, Česlovas

2014-01-01

The analysis of ∼2000 bacterial genomes revealed that they all, without a single exception, encode one or more DNA polymerase III α-subunit (PolIIIα) homologs. Classified into C-family of DNA polymerases they come in two major forms, PolC and DnaE, related by ancient duplication. While PolC represents an evolutionary compact group, DnaE can be further subdivided into at least three groups (DnaE1-3). We performed an extensive analysis of various sequence, structure and surface properties of all four polymerase groups. Our analysis suggests a specific evolutionary pathway leading to PolC and DnaE from the last common ancestor and reveals important differences between extant polymerase groups. Among them, DnaE1 and PolC show the highest conservation of the analyzed properties. DnaE3 polymerases apparently represent an ‘impaired’ version of DnaE1. Nonessential DnaE2 polymerases, typical for oxygen-using bacteria with large GC-rich genomes, have a number of features in common with DnaE3 polymerases. The analysis of polymerase distribution in genomes revealed three major combinations: DnaE1 either alone or accompanied by one or more DnaE2s, PolC + DnaE3 and PolC + DnaE1. The first two combinations are present in Escherichia coli and Bacillus subtilis, respectively. The third one (PolC + DnaE1), found in Clostridia, represents a novel, so far experimentally uncharacterized, set. PMID:24106089

High-fidelity DNA replication in Mycobacterium tuberculosis relies on a trinuclear zinc center.

PubMed

Baños-Mateos, Soledad; van Roon, Anne-Marie M; Lang, Ulla F; Maslen, Sarah L; Skehel, J Mark; Lamers, Meindert H

2017-10-11

High-fidelity DNA replication depends on a proofreading 3'-5' exonuclease that is associated with the replicative DNA polymerase. The replicative DNA polymerase DnaE1 from the major pathogen Mycobacterium tuberculosis (Mtb) uses its intrinsic PHP-exonuclease that is distinct from the canonical DEDD exonucleases found in the Escherichia coli and eukaryotic replisomes. The mechanism of the PHP-exonuclease is not known. Here, we present the crystal structure of the Mtb DnaE1 polymerase. The PHP-exonuclease has a trinuclear zinc center, coordinated by nine conserved residues. Cryo-EM analysis reveals the entry path of the primer strand in the PHP-exonuclease active site. Furthermore, the PHP-exonuclease shows a striking similarity to E. coli endonuclease IV, which provides clues regarding the mechanism of action. Altogether, this work provides important insights into the PHP-exonuclease and reveals unique properties that make it an attractive target for novel anti-mycobacterial drugs.The polymerase and histidinol phosphatase (PHP) domain in the DNA polymerase DnaE1 is essential for mycobacterial high-fidelity DNA replication. Here, the authors determine the DnaE1 crystal structure, which reveals the PHP-exonuclease mechanism that can be exploited for antibiotic development.

A two-dimensional DNA lattice implanted polymer solar cell.

PubMed

Lee, Keun Woo; Kim, Kyung Min; Lee, Junwye; Amin, Rashid; Kim, Byeonghoon; Park, Sung Kye; Lee, Seok Kiu; Park, Sung Ha; Kim, Hyun Jae

2011-09-16

A double crossover tile based artificial two-dimensional (2D) DNA lattice was fabricated and the dry-wet method was introduced to recover an original DNA lattice structure in order to deposit DNA lattices safely on the organic layer without damaging the layer. The DNA lattice was then employed as an electron blocking layer in a polymer solar cell causing an increase of about 10% up to 160% in the power conversion efficiency. Consequently, the resulting solar cell which had an artificial 2D DNA blocking layer showed a significant enhancement in power conversion efficiency compared to conventional polymer solar cells. It should be clear that the artificial DNA nanostructure holds unique physical properties that are extremely attractive for various energy-related and photonic applications.

Direct DNA binding by Brca1.

PubMed

Paull, T T; Cortez, D; Bowers, B; Elledge, S J; Gellert, M

2001-05-22

The tumor suppressor Brca1 plays an important role in protecting mammalian cells against genomic instability, but little is known about its modes of action. In this work we demonstrate that recombinant human Brca1 protein binds strongly to DNA, an activity conferred by a domain in the center of the Brca1 polypeptide. As a result of this binding, Brca1 inhibits the nucleolytic activities of the Mre11/Rad50/Nbs1 complex, an enzyme implicated in numerous aspects of double-strand break repair. Brca1 displays a preference for branched DNA structures and forms protein-DNA complexes cooperatively between multiple DNA strands, but without DNA sequence specificity. This fundamental property of Brca1 may be an important part of its role in DNA repair and transcription.

Comparison of the protein-protein interfaces in the p53-DNA crystal structures: towards elucidation of the biological interface.

PubMed

Ma, Buyong; Pan, Yongping; Gunasekaran, K; Venkataraghavan, R Babu; Levine, Arnold J; Nussinov, Ruth

2005-03-15

p53, the tumor suppressor protein, functions as a dimer of dimers. However, how the tetramer binds to the DNA is still an open question. In the crystal structure, three copies of the p53 monomers (containing chains A, B, and C) were crystallized with the DNA-consensus element. Although the structure provides crucial data on the p53-DNA contacts, the active oligomeric state is unclear because the two dimeric (A-B and B-C) interfaces present in the crystal cannot both exist in the tetramer. Here, we address the question of which of these two dimeric interfaces may be more biologically relevant. We analyze the sequence and structural properties of the p53-p53 dimeric interfaces and carry out extensive molecular dynamics simulations of the crystal structures of the human and mouse p53 dimers. We find that the A-B interface residues are more conserved than those of the B-C. Molecular dynamics simulations show that the A-B interface can provide a stable DNA-binding motif in the dimeric state, unlike B-C. Our results indicate that the interface between chains A-B in the p53-DNA complex constitutes a better candidate for a stable biological interface, whereas the B-C interface is more likely to be due to crystal packing. Thus, they have significant implications toward our understanding of DNA binding by p53 as well as p53-mediated interactions with other proteins.

Disentangling DNA molecules

NASA Astrophysics Data System (ADS)

Vologodskii, Alexander

2016-09-01

The widespread circular form of DNA molecules inside cells creates very serious topological problems during replication. Due to the helical structure of the double helix the parental strands of circular DNA form a link of very high order, and yet they have to be unlinked before the cell division. DNA topoisomerases, the enzymes that catalyze passing of one DNA segment through another, solve this problem in principle. However, it is very difficult to remove all entanglements between the replicated DNA molecules due to huge length of DNA comparing to the cell size. One strategy that nature uses to overcome this problem is to create the topoisomerases that can dramatically reduce the fraction of linked circular DNA molecules relative to the corresponding fraction at thermodynamic equilibrium. This striking property of the enzymes means that the enzymes that interact with DNA only locally can access their topology, a global property of circular DNA molecules. This review considers the experimental studies of the phenomenon and analyzes the theoretical models that have been suggested in attempts to explain it. We describe here how various models of enzyme action can be investigated computationally. There is no doubt at the moment that we understand basic principles governing enzyme action. Still, there are essential quantitative discrepancies between the experimental data and the theoretical predictions. We consider how these discrepancies can be overcome.

Synthesis of mouse centromere-targeted polyamides and physico-chemical studies of their interaction with the target double-stranded DNA.

PubMed

Nozeret, Karine; Bonan, Marc; Yarmoluk, Serguiy M; Novopashina, Darya S; Boutorine, Alexandre S

2015-09-01

Synthetic minor groove-binding pyrrole-imidazole polyamides labeled by fluorophores are promising candidates for fluorescence imaging of double-stranded DNA in isolated chromosomes or fixed and living cells. We synthesized nine hairpin and two head-to-head tandem polyamides targeting repeated sequences from mouse major satellites. Their interaction with synthetic target dsDNA has been studied by physico-chemical methods in vitro before and after coupling to various fluorophores. Great variability in affinities and fluorescence properties reveals a conclusion that these properties do not only rely on recognition rules, but also on other known and unknown structural factors. Individual testing of each probe is needed before cellular applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

Mechanical Properties of Transcription

NASA Astrophysics Data System (ADS)

Sevier, Stuart A.; Levine, Herbert

2017-06-01

The mechanical properties of transcription have recently been shown to play a central role in gene expression. However, a full physical characterization of this central biological process is lacking. In this Letter, we introduce a simple description of the basic physical elements of transcription where RNA elongation, RNA polymerase rotation, and DNA supercoiling are coupled. The resulting framework describes the relative amount of RNA polymerase rotation and DNA supercoiling that occurs during RNA elongation. Asymptotic behavior is derived and can be used to experimentally extract unknown mechanical parameters of transcription. Mechanical limits to transcription are incorporated through the addition of a DNA supercoiling-dependent RNA polymerase velocity. This addition can lead to transcriptional stalling and resulting implications for gene expression, chromatin structure and genome organization are discussed.

The centromeric nucleosome-like CENP–T–W–S–X complex induces positive supercoils into DNA

PubMed Central

Takeuchi, Kozo; Nishino, Tatsuya; Mayanagi, Kouta; Horikoshi, Naoki; Osakabe, Akihisa; Tachiwana, Hiroaki; Hori, Tetsuya; Kurumizaka, Hitoshi; Fukagawa, Tatsuo

2014-01-01

The centromere is a specific genomic region upon which the kinetochore is formed to attach to spindle microtubules for faithful chromosome segregation. To distinguish this chromosomal region from other genomic loci, the centromere contains a specific chromatin structure including specialized nucleosomes containing the histone H3 variant CENP–A. In addition to CENP–A nucleosomes, we have found that centromeres contain a nucleosome-like structure comprised of the histone-fold CENP–T–W–S–X complex. However, it is unclear how the CENP–T–W–S–X complex associates with centromere chromatin. Here, we demonstrate that the CENP–T–W–S–X complex binds preferentially to ∼100 bp of linker DNA rather than nucleosome-bound DNA. In addition, we find that the CENP–T–W–S–X complex primarily binds to DNA as a (CENP–T–W–S–X)2 structure. Interestingly, in contrast to canonical nucleosomes that negatively supercoil DNA, the CENP–T–W–S–X complex induces positive DNA supercoils. We found that the DNA-binding regions in CENP–T or CENP–W, but not CENP–S or CENP–X, are required for this positive supercoiling activity and the kinetochore targeting of the CENP–T–W–S–X complex. In summary, our work reveals the structural features and properties of the CENP–T–W–S–X complex for its localization to centromeres. PMID:24234442

DNA nanoparticles with core-shell morphology.

PubMed

Chandran, Preethi L; Dimitriadis, Emilios K; Lisziewicz, Julianna; Speransky, Vlad; Horkay, Ferenc

2014-10-14

Mannobiose-modified polyethylenimines (PEI) are used in gene therapy to generate nanoparticles of DNA that can be targeted to the antigen-presenting cells of the immune system. We report that the sugar modification alters the DNA organization within the nanoparticles from homogenous to shell-like packing. The depth-dependent packing of DNA within the nanoparticles was probed using AFM nano-indentation. Unmodified PEI-DNA nanoparticles display linear elastic properties and depth-independent mechanics, characteristic of homogenous materials. Mannobiose-modified nanoparticles, however, showed distinct force regimes that were dependent on indentation depth, with 'buckling'-like response that is reproducible and not due to particle failure. By comparison with theoretical studies of spherical shell mechanics, the structure of mannobiosylated particles was deduced to be a thin shell with wall thickness in the order of few nanometers, and a fluid-filled core. The shell-core structure is also consistent with observations of nanoparticle denting in altered solution conditions, with measurements of nanoparticle water content from AFM images, and with images of DNA distribution in Transmission Electron Microscopy.

Structure-based discovery of NANOG variant with enhanced properties to promote self-renewal and reprogramming of pluripotent stem cells

DOE PAGES

Hayashi, Yohei; Caboni, Laura; Das, Debanu; ...

2015-03-30

NANOG (from Irish mythology Tír na nÓg) transcription factor plays a central role in maintaining pluripotency, cooperating with OCT4 (also known as POU5F1 or OCT3/4), SOX2, and other pluripotency factors. Although the physiological roles of the NANOG protein have been extensively explored, biochemical and biophysical properties in relation to its structural analysis are poorly understood. Here we determined the crystal structure of the human NANOG homeodomain (hNANOG HD) bound to an OCT4 promoter DNA, which revealed amino acid residues involved in DNA recognition that are likely to be functionally important. We generated a series of hNANOG HD alanine substitution mutantsmore » based on the protein–DNA interaction and evolutionary conservation and determined their biological activities. Some mutant proteins were less stable, resulting in loss or decreased affinity for DNA binding. Overexpression of the orthologous mouse NANOG (mNANOG) mutants failed to maintain self-renewal of mouse embryonic stem cells without leukemia inhibitory factor. These results suggest that these residues are critical for NANOG transcriptional activity. Interestingly, one mutant, hNANOG L122A, conversely enhanced protein stability and DNA-binding affinity. The mNANOG L122A, when overexpressed in mouse embryonic stem cells, maintained their expression of self-renewal markers even when retinoic acid was added to forcibly drive differentiation. When overexpressed in epiblast stem cells or human induced pluripotent stem cells, the L122A mutants enhanced reprogramming into ground-state pluripotency. These findings indicate that structural and biophysical information on key transcriptional factors provides insights into the manipulation of stem cell behaviors and a framework for rational protein engineering.« less

Structure-based discovery of NANOG variant with enhanced properties to promote self-renewal and reprogramming of pluripotent stem cells

PubMed Central

Hayashi, Yohei; Caboni, Laura; Das, Debanu; Yumoto, Fumiaki; Clayton, Thomas; Deller, Marc C.; Nguyen, Phuong; Farr, Carol L.; Chiu, Hsiu-Ju; Miller, Mitchell D.; Elsliger, Marc-André; Deacon, Ashley M.; Godzik, Adam; Lesley, Scott A.; Tomoda, Kiichiro; Conklin, Bruce R.; Wilson, Ian A.; Yamanaka, Shinya; Fletterick, Robert J.

2015-01-01

NANOG (from Irish mythology Tír na nÓg) transcription factor plays a central role in maintaining pluripotency, cooperating with OCT4 (also known as POU5F1 or OCT3/4), SOX2, and other pluripotency factors. Although the physiological roles of the NANOG protein have been extensively explored, biochemical and biophysical properties in relation to its structural analysis are poorly understood. Here we determined the crystal structure of the human NANOG homeodomain (hNANOG HD) bound to an OCT4 promoter DNA, which revealed amino acid residues involved in DNA recognition that are likely to be functionally important. We generated a series of hNANOG HD alanine substitution mutants based on the protein–DNA interaction and evolutionary conservation and determined their biological activities. Some mutant proteins were less stable, resulting in loss or decreased affinity for DNA binding. Overexpression of the orthologous mouse NANOG (mNANOG) mutants failed to maintain self-renewal of mouse embryonic stem cells without leukemia inhibitory factor. These results suggest that these residues are critical for NANOG transcriptional activity. Interestingly, one mutant, hNANOG L122A, conversely enhanced protein stability and DNA-binding affinity. The mNANOG L122A, when overexpressed in mouse embryonic stem cells, maintained their expression of self-renewal markers even when retinoic acid was added to forcibly drive differentiation. When overexpressed in epiblast stem cells or human induced pluripotent stem cells, the L122A mutants enhanced reprogramming into ground-state pluripotency. These findings demonstrate that structural and biophysical information on key transcriptional factors provides insights into the manipulation of stem cell behaviors and a framework for rational protein engineering. PMID:25825768

Structure-based discovery of NANOG variant with enhanced properties to promote self-renewal and reprogramming of pluripotent stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hayashi, Yohei; Caboni, Laura; Das, Debanu

NANOG (from Irish mythology Tír na nÓg) transcription factor plays a central role in maintaining pluripotency, cooperating with OCT4 (also known as POU5F1 or OCT3/4), SOX2, and other pluripotency factors. Although the physiological roles of the NANOG protein have been extensively explored, biochemical and biophysical properties in relation to its structural analysis are poorly understood. Here we determined the crystal structure of the human NANOG homeodomain (hNANOG HD) bound to an OCT4 promoter DNA, which revealed amino acid residues involved in DNA recognition that are likely to be functionally important. We generated a series of hNANOG HD alanine substitution mutantsmore » based on the protein–DNA interaction and evolutionary conservation and determined their biological activities. Some mutant proteins were less stable, resulting in loss or decreased affinity for DNA binding. Overexpression of the orthologous mouse NANOG (mNANOG) mutants failed to maintain self-renewal of mouse embryonic stem cells without leukemia inhibitory factor. These results suggest that these residues are critical for NANOG transcriptional activity. Interestingly, one mutant, hNANOG L122A, conversely enhanced protein stability and DNA-binding affinity. The mNANOG L122A, when overexpressed in mouse embryonic stem cells, maintained their expression of self-renewal markers even when retinoic acid was added to forcibly drive differentiation. When overexpressed in epiblast stem cells or human induced pluripotent stem cells, the L122A mutants enhanced reprogramming into ground-state pluripotency. These findings indicate that structural and biophysical information on key transcriptional factors provides insights into the manipulation of stem cell behaviors and a framework for rational protein engineering.« less

An unconventional family 1 uracil DNA glycosylase in Nitratifractor salsuginis.

PubMed

Li, Jing; Chen, Ran; Yang, Ye; Zhang, Zhemin; Fang, Guang-Chen; Xie, Wei; Cao, Weiguo

2017-12-01

The uracil DNA glycosylase superfamily consists of at least six families with a diverse specificity toward DNA base damage. Family 1 uracil N-glycosylase (UNG) exhibits exclusive specificity on uracil-containing DNA. Here, we report a family 1 UNG homolog from Nitratifractor salsuginis with distinct biochemical features that differentiate it from conventional family 1 UNGs. Globally, the crystal structure of N. salsuginisUNG shows a few additional secondary structural elements. Biochemical and enzyme kinetic analysis, coupled with structural determination, molecular modeling, and molecular dynamics simulations, shows that N. salsuginisUNG contains a salt bridge network that plays an important role in DNA backbone interactions. Disruption of the amino acid residues involved in the salt bridges greatly impedes the enzymatic activity. A tyrosine residue in motif 1 (GQDPY) is one of the distinct sequence features setting family 1 UNG apart from other families. The crystal structure of Y81G mutant indicates that several subtle changes may account for its inactivity. Unlike the conventional family 1 UNG enzymes, N. salsuginisUNG is not inhibited by Ugi, a potent inhibitor specific for family 1 UNG. This study underscores the diversity of paths that a uracil DNA glycosylase may take to acquire its unique structural and biochemical properties during evolution. Structure data are available in the PDB under accession numbers 5X3G and 5X3H. © 2017 Federation of European Biochemical Societies.

Structural and Functional Studies of H. seropedicae RecA Protein – Insights into the Polymerization of RecA Protein as Nucleoprotein Filament

DOE Office of Scientific and Technical Information (OSTI.GOV)

Leite, Wellington C.; Galvão, Carolina W.; Saab, Sérgio C.

The bacterial RecA protein plays a role in the complex system of DNA damage repair. Here, we report the functional and structural characterization of the Herbaspirillum seropedicae RecA protein (HsRecA). HsRecA protein is more efficient at displacing SSB protein from ssDNA than Escherichia coli RecA protein. HsRecA also promotes DNA strand exchange more efficiently. The three dimensional structure of HsRecA-ADP/ATP complex has been solved to 1.7 Å resolution. HsRecA protein contains a small N-terminal domain, a central core ATPase domain and a large C-terminal domain, that are similar to homologous bacterial RecA proteins. Comparative structural analysis showed that the N-terminalmore » polymerization motif of archaeal and eukaryotic RecA family proteins are also present in bacterial RecAs. Reconstruction of electrostatic potential from the hexameric structure of HsRecA-ADP/ATP revealed a high positive charge along the inner side, where ssDNA is bound inside the filament. The properties of this surface may explain the greater capacity of HsRecA protein to bind ssDNA, forming a contiguous nucleoprotein filament, displace SSB and promote DNA exchange relative to EcRecA. In conclusion, our functional and structural analyses provide insight into the molecular mechanisms of polymerization of bacterial RecA as a helical nucleoprotein filament.« less

Structural and Functional Studies of H. seropedicae RecA Protein – Insights into the Polymerization of RecA Protein as Nucleoprotein Filament

PubMed Central

Galvão, Carolina W.; Saab, Sérgio C.; Iulek, Jorge; Etto, Rafael M.; Steffens, Maria B. R.; Chitteni-Pattu, Sindhu; Stanage, Tyler; Keck, James L.; Cox, Michael M.

2016-01-01

The bacterial RecA protein plays a role in the complex system of DNA damage repair. Here, we report the functional and structural characterization of the Herbaspirillum seropedicae RecA protein (HsRecA). HsRecA protein is more efficient at displacing SSB protein from ssDNA than Escherichia coli RecA protein. HsRecA also promotes DNA strand exchange more efficiently. The three dimensional structure of HsRecA-ADP/ATP complex has been solved to 1.7 Å resolution. HsRecA protein contains a small N-terminal domain, a central core ATPase domain and a large C-terminal domain, that are similar to homologous bacterial RecA proteins. Comparative structural analysis showed that the N-terminal polymerization motif of archaeal and eukaryotic RecA family proteins are also present in bacterial RecAs. Reconstruction of electrostatic potential from the hexameric structure of HsRecA-ADP/ATP revealed a high positive charge along the inner side, where ssDNA is bound inside the filament. The properties of this surface may explain the greater capacity of HsRecA protein to bind ssDNA, forming a contiguous nucleoprotein filament, displace SSB and promote DNA exchange relative to EcRecA. Our functional and structural analyses provide insight into the molecular mechanisms of polymerization of bacterial RecA as a helical nucleoprotein filament. PMID:27447485

G-quadruplexes Significantly Stimulate Pif1 Helicase-catalyzed Duplex DNA Unwinding*

PubMed Central

Duan, Xiao-Lei; Liu, Na-Nv; Yang, Yan-Tao; Li, Hai-Hong; Li, Ming; Dou, Shuo-Xing; Xi, Xu-Guang

2015-01-01

The evolutionarily conserved G-quadruplexes (G4s) are faithfully inherited and serve a variety of cellular functions such as telomere maintenance, gene regulation, DNA replication initiation, and epigenetic regulation. Different from the Watson-Crick base-pairing found in duplex DNA, G4s are formed via Hoogsteen base pairing and are very stable and compact DNA structures. Failure of untangling them in the cell impedes DNA-based transactions and leads to genome instability. Cells have evolved highly specific helicases to resolve G4 structures. We used a recombinant nuclear form of Saccharomyces cerevisiae Pif1 to characterize Pif1-mediated DNA unwinding with a substrate mimicking an ongoing lagging strand synthesis stalled by G4s, which resembles a replication origin and a G4-structured flap in Okazaki fragment maturation. We find that the presence of G4 may greatly stimulate the Pif1 helicase to unwind duplex DNA. Further studies reveal that this stimulation results from G4-enhanced Pif1 dimerization, which is required for duplex DNA unwinding. This finding provides new insights into the properties and functions of G4s. We discuss the observed activation phenomenon in relation to the possible regulatory role of G4s in the rapid rescue of the stalled lagging strand synthesis by helping the replicator recognize and activate the replication origin as well as by quickly removing the G4-structured flap during Okazaki fragment maturation. PMID:25627683

Coarse-grained simulation of DNA using LAMMPS : An implementation of the oxDNA model and its applications.

PubMed

Henrich, Oliver; Gutiérrez Fosado, Yair Augusto; Curk, Tine; Ouldridge, Thomas E

2018-05-10

During the last decade coarse-grained nucleotide models have emerged that allow us to study DNA and RNA on unprecedented time and length scales. Among them is oxDNA, a coarse-grained, sequence-specific model that captures the hybridisation transition of DNA and many structural properties of single- and double-stranded DNA. oxDNA was previously only available as standalone software, but has now been implemented into the popular LAMMPS molecular dynamics code. This article describes the new implementation and analyses its parallel performance. Practical applications are presented that focus on single-stranded DNA, an area of research which has been so far under-investigated. The LAMMPS implementation of oxDNA lowers the entry barrier for using the oxDNA model significantly, facilitates future code development and interfacing with existing LAMMPS functionality as well as other coarse-grained and atomistic DNA models.

Defined-size DNA triple crossover construct for molecular electronics: modification, positioning and conductance properties.

PubMed

Linko, Veikko; Leppiniemi, Jenni; Paasonen, Seppo-Tapio; Hytönen, Vesa P; Toppari, J Jussi

2011-07-08

We present a novel, defined-size, small and rigid DNA template, a so-called B-A-B complex, based on DNA triple crossover motifs (TX tiles), which can be utilized in molecular scale patterning for nanoelectronics, plasmonics and sensing applications. The feasibility of the designed construct is demonstrated by functionalizing the TX tiles with one biotin-triethylene glycol (TEG) and efficiently decorating them with streptavidin, and furthermore by positioning and anchoring single thiol-modified B-A-B complexes to certain locations on a chip via dielectrophoretic trapping. Finally, we characterize the conductance properties of the non-functionalized construct, first by measuring DC conductivity and second by utilizing AC impedance spectroscopy in order to describe the conductivity mechanism of a single B-A-B complex using a detailed equivalent circuit model. This analysis also reveals further information about the conductivity of DNA structures in general.

THE EFFECT OF CHLORINATION OF NUCLEOTIDE BASES ON THE CONFORMATIONAL PROPERTIES OF THYMIDINE MONOPHOSPHATE.

PubMed

Mukhina, T M; Nikolaienko, T Yu

2015-01-01

Recent studies on Escherichia coli bacteria cultivation, in which DNA thymine was replaced with 5-chlorouracil have refreshed the problem of understanding the changes to physical properties of DNA monomers resultant from chemical modifications. These studies have shown that the replacement did not affect the normal activities and division of the bacteria, but has significantly reduced its life span. In this paper a comparative analysis was carried out by the methods of computational experiment of a set of 687 possible conformers of natural monomeric DNA unit (2'-deoxyribonucleotide thymidine monophosphate) and 660 conformers of 5-chloro-2'-deoxyuridine monophosphate - a similar molecules in which the natural nitrogenous base thymine is substituted with 5-chlorouracil. Structures of stable conformers of the modified deoxyribonucleotide have been obtained and physical factors, which determine their variation from the conformers of the unmodified molecule have been analyzed. A comparative analysis of the elastic properties of conformers of investigated molecules and non-covalent interactions present in them was conducted. The results can be usedfor planning experiments on synthesis of artficial DNA suitable for incorporation into living organisms.

Analysis of informational redundancy in the protein-assembling machinery

NASA Astrophysics Data System (ADS)

Berkovich, Simon

2004-03-01

Entropy analysis of the DNA structure does not reveal a significant departure from randomness indicating lack of informational redundancy. This signifies the absence of a hidden meaning in the genome text and supports the 'barcode' interpretation of DNA given in [1]. Lack of informational redundancy is a characteristic property of an identification label rather than of a message of instructions. Yet randomness of DNA has to induce non-random structures of the proteins. Protein synthesis is a two-step process: transcription into RNA with gene splicing and formation a structure of amino acids. Entropy estimations, performed by A. Djebbari, show typical values of redundancy of the biomolecules along these pathways: DNA gene 4proteins 15-40in gene expression, the RNA copy carries the same information as the original DNA template. Randomness is essentially eliminated only at the step of the protein creation by a degenerate code. According to [1], the significance of the substitution of U for T with a subsequent gene splicing is that these transformations result in a different pattern of RNA oscillations, so the vital DNA communications are protected against extraneous noise coming from the protein making activities. 1. S. Berkovich, "On the 'barcode' functionality of DNA, or the Phenomenon of Life in the Physical Universe", Dorrance Publishing Co., Pittsburgh, 2003

Effect of Fe3+ on Curcumin-DNA Complex Studied by FT-Raman Spectroscopy

NASA Astrophysics Data System (ADS)

Senthil, K.; Sarojini, R.

2008-11-01

Curcumin is a non toxic and natural antioxidant. The interaction of curcumin with DNA is widely used in clinical treatment of a variety of different forms of cancer, but its antioxidant property turns into proxidant in the presence of high Fe3+ concentrations. However, the relevance of the pro-oxidant nature of curcumin at molecular level is not clearly determined. Thus, in the present study the interaction of Fe(III) on Curcumin DNA complexes were investigated at physiological pH with Fe(III)/drug/DNA (Phosphate) molar ratios (r) 1:10:50, 1:5:25, 1:2:10 and 1:1:5. FT-Raman Spectroscopy was used to establish correlation between spectral changes and drug binding mode, sequence selectivity, DNA Conformation and Structural properties of Fe(III)/drug/DNA complexes in aqueous solution. Spectroscopic results showed that the major spectral changes were observed at 688 cm-1(G), 835 cm-1 (O-P-O), 1092 cm-1 (PO2-), 1485 cm-1 (G,A,T) and 1683 cm-1 (T), 1723 cm-1 (G) in Fe(III)/drug/DNA complex indicating affinity of Fe(III)/drug with the phosphate and DNA base pairs. The present result showed that the combination of Fe-curcumin induced significant DNA damage in a concentration dependent manner.

DNA Nanotubes for NMR Structure Determination of Membrane Proteins

PubMed Central

Bellot, Gaëtan; McClintock, Mark A.; Chou, James J; Shih, William M.

2013-01-01

Structure determination of integral membrane proteins by solution NMR represents one of the most important challenges of structural biology. A Residual-Dipolar-Coupling-based refinement approach can be used to solve the structure of membrane proteins up to 40 kDa in size, however, a weak-alignment medium that is detergent-resistant is required. Previously, availability of media suitable for weak alignment of membrane proteins was severely limited. We describe here a protocol for robust, large-scale synthesis of detergent-resistant DNA nanotubes that can be assembled into dilute liquid crystals for application as weak-alignment media in solution NMR structure determination of membrane proteins in detergent micelles. The DNA nanotubes are heterodimers of 400nm-long six-helix bundles each self-assembled from a M13-based p7308 scaffold strand and >170 short oligonucleotide staple strands. Compatibility with proteins bearing considerable positive charge as well as modulation of molecular alignment, towards collection of linearly independent restraints, can be introduced by reducing the negative charge of DNA nanotubes via counter ions and small DNA binding molecules. This detergent-resistant liquid-crystal media offers a number of properties conducive for membrane protein alignment, including high-yield production, thermal stability, buffer compatibility, and structural programmability. Production of sufficient nanotubes for 4–5 NMR experiments can be completed in one week by a single individual. PMID:23518667

A Novel DNA Binding Mechanism for maf Basic Region-Leucine Zipper Factors Inferred from a MafA-DNA Complex Structure and Binding Specificities

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu, Xun; Guanga, Gerald P; Wan, Cheng

2012-11-13

MafA is a proto-oncoprotein and is critical for insulin gene expression in pancreatic β-cells. Maf proteins belong to the AP1 superfamily of basic region-leucine zipper (bZIP) transcription factors. Residues in the basic helix and an ancillary N-terminal domain, the Extended Homology Region (EHR), endow maf proteins with unique DNA binding properties: binding a 13 bp consensus site consisting of a core AP1 site (TGACTCA) flanked by TGC sequences and binding DNA stably as monomers. To further characterize maf DNA binding, we determined the structure of a MafA–DNA complex. MafA forms base-specific hydrogen bonds with the flanking G –5C –4 andmore » central C 0/G 0 bases, but not with the core-TGA bases. However, in vitro binding studies utilizing a pulse–chase electrophoretic mobility shift assay protocol revealed that mutating either the core-TGA or flanking-TGC bases dramatically increases the binding off rate. Comparing the known maf structures, we propose that DNA binding specificity results from positioning the basic helix through unique phosphate contacts. The EHR does not contact DNA directly but stabilizes DNA binding by contacting the basic helix. Collectively, these results suggest a novel multistep DNA binding process involving a conformational change from contacting the core-TGA to contacting the flanking-TGC bases.« less

Noncanonical structures and their thermodynamics of DNA and RNA under molecular crowding: beyond the Watson-Crick double helix.

PubMed

Sugimoto, Naoki

2014-01-01

How does molecular crowding affect the stability of nucleic acid structures inside cells? Water is the major solvent component in living cells, and the properties of water in the highly crowded media inside cells differ from that in buffered solution. As it is difficult to measure the thermodynamic behavior of nucleic acids in cells directly and quantitatively, we recently developed a cell-mimicking system using cosolutes as crowding reagents. The influences of molecular crowding on the structures and thermodynamics of various nucleic acid sequences have been reported. In this chapter, we discuss how the structures and thermodynamic properties of nucleic acids differ under various conditions such as highly crowded environments, compartment environments, and in the presence of ionic liquids, and the major determinants of the crowding effects on nucleic acids are discussed. The effects of molecular crowding on the activities of ribozymes and riboswitches on noncanonical structures of DNA- and RNA-like quadruplexes that play important roles in transcription and translation are also described. © 2014 Elsevier Inc. All rights reserved.

Static and Dynamic Properties of DNA Confined in Nanochannels

NASA Astrophysics Data System (ADS)

Gupta, Damini

Next-generation sequencing (NGS) techniques have considerably reduced the cost of high-throughput DNA sequencing. However, it is challenging to detect large-scale genomic variations by NGS due to short read lengths. Genome mapping can easily detect large-scale structural variations because it operates on extremely large intact molecules of DNA with adequate resolution. One of the promising methods of genome mapping is based on confining large DNA molecules inside a nanochannel whose cross-sectional dimensions are approximately 50 nm. Even though this genome mapping technology has been commercialized, the current understanding of the polymer physics of DNA in nanochannel confinement is based on theories and lacks much needed experimental support. The results of this dissertation are aimed at providing a detailed experimental understanding of equilibrium properties of nanochannel-confined DNA molecules. The results are divided into three parts. In first part, we evaluate the role of channel shape on thermodynamic properties of channel confined DNA molecules using a combination of fluorescence microscopy and simulations. Specifically, we show that high aspect ratio of rectangular channels significantly alters the chain statistics as compared to an equivalent square channel with same cross-sectional area. In the second part, we present experimental evidence that weak excluded volume effects arise in DNA nanochannel confinement, which form the physical basis for the extended de Gennes regime. We also show how confinement spectroscopy and simulations can be combined to reduce molecular weight dispersity effects arising from shearing, photo-cleavage, and nonuniform staining of DNA. Finally, the third part of the thesis concerns the dynamic properties of nanochannel confined DNA. We directly measure the center-of-mass diffusivity of single DNA molecules in confinement and show that that it is necessary to modify the classical results of de Gennes to account for local chain stiffness of DNA in order to explain the experimental results. In the end, we believe that our findings from the experimental test of the phase diagram for channel-confined DNA, with careful control over molecular weight dispersity, channel geometry, and electrostatic interactions, will provide a firm foundation for the emerging genome mapping technology.

Unveiling the Mode of Interaction of Berberine Alkaloid in Different Supramolecular Confined Environments: Interplay of Surface Charge between Nano-Confined Charged Layer and DNA.

PubMed

Kundu, Niloy; Roy, Arpita; Banik, Debasis; Sarkar, Nilmoni

2016-02-18

In this Article, we demonstrate a detailed characterization of binding interaction of berberine chloride (BBCl) with calf-thymus DNA (CT-DNA) in buffer solution as well as in two differently charged reverse micelles (RMs). The photophyscial properties of this alkaloid have been modulated within these microheterogeneous bioassemblies. The mode of binding of this alkaloid with DNA is of debate to date. However, fluorescence spectroscopic measurements, circular dichroism (CD) measurement, and temperature-dependent study unambiguously establish that BBCl partially intercalates into the DNA base pairs. The nonplanarity imposed by partial saturation in their structure causes the nonclassical types of intercalation into DNA. Besides the intercalation, electrostatic interactions also play a significant role in the binding between BBCl and DNA. DNA structure turns into a condensed form after encapsulation into RMs, which is followed by the CD spectra and microscopy study. The probe location and dynamics in the nanopool of the RMs depended on the electrostatic interaction between the charged surfactants and cationic berberine. The structural alteration of CT-DNA from B form to condensed form and the interplay of surface charge between RMs and DNA determine the interaction between the alkaloid and DNA in RMs. Time-resolved study and fluorescence anisotropy measurements successfully provide the binding interaction of BBCl in the nanopool of the RMs in the absence and in the presence of DNA. This study motivates us to judge further the potential applicability of this alkaloid in other biological systems or other biomimicking organized assemblies.

Binding of anticancer drug daunomycin to a TGGGGT G-quadruplex DNA probed by all-atom molecular dynamics simulations: additional pure groove binding mode and implications on designing more selective G-quadruplex ligands.

PubMed

Shen, Zhanhang; Mulholland, Kelly A; Zheng, Yujun; Wu, Chun

2017-09-01

DNA G-quadruplex structures are emerging cancer-specific targets for chemotherapeutics. Ligands that bind to and stabilize DNA G-quadruplexes have the potential to be anti-cancer drugs. Lack of binding selectivity to DNA G-quadruplex over DNA duplex remains a major challenge when attempting to develop G-quadruplex ligands into successful anti-cancer drugs. Thorough understanding of the binding nature of existing non-selective ligands that bind to both DNA quadruplex and DNA duplex will help to address this challenge. Daunomycin and doxorubicin, two commonly used anticancer drugs, are examples of non-selective DNA ligands. In this study, we extended our early all-atom binding simulation studies between doxorubicin and a DNA duplex (d(CGATCG) 2 ) to probe the binding between daunomycin and a parallel DNA quadruplex (d(TGGGGT) 4 ) and DNA duplex. In addition to the end stacking mode, which mimics the mode in the crystal structure, a pure groove binding mode was observed in our free binding simulations. The dynamic and energetic properties of these two binding modes are thoroughly examined, and a detailed comparison is made between DNA quadruplex binding modes and DNA duplex binding modes. Implications on the design of more selective DNA quadruplex ligands are also discussed. Graphical abstract Top stacking and groov binding modes from the MD simulations.

(CAG)(n)-hairpin DNA binds to Msh2-Msh3 and changes properties of mismatch recognition.

PubMed

Owen, Barbara A L; Yang, Zungyoon; Lai, Maoyi; Gajec, Maciej; Gajek, Maciez; Badger, John D; Hayes, Jeffrey J; Edelmann, Winfried; Kucherlapati, Raju; Wilson, Teresa M; McMurray, Cynthia T

2005-08-01

Cells have evolved sophisticated DNA repair systems to correct damaged DNA. However, the human DNA mismatch repair protein Msh2-Msh3 is involved in the process of trinucleotide (CNG) DNA expansion rather than repair. Using purified protein and synthetic DNA substrates, we show that Msh2-Msh3 binds to CAG-hairpin DNA, a prime candidate for an expansion intermediate. CAG-hairpin binding inhibits the ATPase activity of Msh2-Msh3 and alters both nucleotide (ADP and ATP) affinity and binding interfaces between protein and DNA. These changes in Msh2-Msh3 function depend on the presence of A.A mispaired bases in the stem of the hairpin and on the hairpin DNA structure per se. These studies identify critical functional defects in the Msh2-Msh3-CAG hairpin complex that could misdirect the DNA repair process.

Precision polymers and 3D DNA nanostructures: emergent assemblies from new parameter space.

PubMed

Serpell, Christopher J; Edwardson, Thomas G W; Chidchob, Pongphak; Carneiro, Karina M M; Sleiman, Hanadi F

2014-11-05

Polymer self-assembly and DNA nanotechnology have both proved to be powerful nanoscale techniques. To date, most attempts to merge the fields have been limited to placing linear DNA segments within a polydisperse block copolymer. Here we show that, by using hydrophobic polymers of a precisely predetermined length conjugated to DNA strands, and addressable 3D DNA prisms, we are able to effect the formation of unprecedented monodisperse quantized superstructures. The structure and properties of larger micelles-of-prisms were probed in depth, revealing their ability to participate in controlled release of their constituent nanostructures, and template light-harvesting energy transfer cascades, mediated through both the addressability of DNA and the controlled aggregation of the polymers.

Modulation of DNA binding by gene-specific transcription factors.

PubMed

Schleif, Robert F

2013-10-01

The transcription of many genes, particularly in prokaryotes, is controlled by transcription factors whose activity can be modulated by controlling their DNA binding affinity. Understanding the molecular mechanisms by which DNA binding affinity is regulated is important, but because forming definitive conclusions usually requires detailed structural information in combination with data from extensive biophysical, biochemical, and sometimes genetic experiments, little is truly understood about this topic. This review describes the biological requirements placed upon DNA binding transcription factors and their consequent properties, particularly the ways that DNA binding affinity can be modulated and methods for its study. What is known and not known about the mechanisms modulating the DNA binding affinity of a number of prokaryotic transcription factors, including CAP and lac repressor, is provided.

Interaction of vasicine with calf thymus DNA: Molecular docking, spectroscopic and differential scanning calorimetric insights

NASA Astrophysics Data System (ADS)

R. S., Sai Murali; R. S., Sai Siddhardha; Rajesh Babu, D.; Venketesh, S.; Basavaraju, R.; Nageswara Rao, G.

2017-06-01

The present study brings out the interaction between vasicine, an alkaloid and Adhatoda vasica Nees with double stranded DNA. The physico-chemical interaction between small molecules and nucleic acids is a major area of focus in screening drugs against various cancers. Molecular probing in our study using Molecular Operating Environment (MOE) has revealed interaction of vasicine with DNA double helix. Here we report the interaction of vasicine with Calf thymus DNA. We present for the first time the results obtained from UV-visible, fluorescence spectroscopic and differential scanning calorimetric techniques that suggest a moderate to strong electrostatic, hydrophobic and van der Waals interactions mediating the DNA binding properties of vasicine, leading to disruption of DNA secondary structure.

Spiroiminodihydantoin lesions derived from guanine oxidation: structures, energetics, and functional implications.

PubMed

Jia, Lei; Shafirovich, Vladimir; Shapiro, Robert; Geacintov, Nicholas E; Broyde, Suse

2005-04-26

Reactive oxygen species present in the cell generate DNA damage. One of the major oxidation products of guanine in DNA, 8-oxo-7,8-dihydroguanine, formed by loss of two electrons, is among the most extensively studied base lesions. The further removal of two electrons from this product can yield spiroiminodihydantoin (Sp) R and S stereoisomers. Both in vitro and in vivo experiments have shown that the Sp stereoisomers are highly mutagenic, causing G --> T and G --> C transversions. Hence, they are of interest as examples of endogenous DNA damage that may initiate cancer. To interpret the mutagenic properties of the Sp lesions, an understanding of their structural properties is needed. To elucidate these structural effects, we have carried out computational investigations at the level of the Sp-modified base and nucleoside. At the base level, quantum mechanical geometry optimization studies have revealed exact mirror image symmetry of the R and S stereoisomers, with a near-perpendicular geometry of the two rings. At the nucleoside level, an extensive survey of the potential energy surface by molecular mechanics calculations using AMBER has provided three-dimensional potential energy maps. These maps reveal that the range and flexibility of the glycosidic torsion angles are significantly more restricted in both stereoisomeric adducts than in unmodified 2'-deoxyguanosine. The structural and energetic results suggest that the unusual geometric, steric, and hydrogen bonding properties of these lesions underlie their mutagenicity. In addition, stereoisomer-specific differences indicate the possibility that their processing by cellular replication and repair enzymes may be differentially affected by their absolute configuration.

Microwave-Induced Inactivation of DNA-Based Hybrid Catalyst in Asymmetric Catalysis

PubMed Central

Zhao, Hua; Shen, Kai

2015-01-01

DNA-based hybrid catalysts have gained strong interests in asymmetric reactions. However, to maintain the high enantioselectivity, these reactions are usually conducted at relatively low temperatures (e.g. < 5 °C) for 2–3 days. Aiming to improve the reaction’s turnover rate, we evaluated microwave irradiation with simultaneous cooling as potential energy source since this method has been widely used to accelerate various chemical and enzymatic reactions. However, our data indicated that microwave irradiation induced an inactivation of DNA-based hybrid catalyst even at low temperatures (such as 5 °C). Circular dichroism (CD) spectra and gel electrophoresis of DNA suggest that microwave exposure degrades DNA molecules and disrupts DNA double-stranded structures, causing changes of DNA–metal ligand binding properties and thus poor DNA catalytic performance. PMID:26712696

DNA-controlled assembly of a NaTl lattice structure from gold nanoparticles and protein nanoparticles

NASA Astrophysics Data System (ADS)

Cigler, Petr; Lytton-Jean, Abigail K. R.; Anderson, Daniel G.; Finn, M. G.; Park, Sung Yong

2010-11-01

The formation of diamond structures from tailorable building blocks is an important goal in colloidal crystallization because the non-compact diamond lattice is an essential component of photonic crystals for the visible-light range. However, designing nanoparticle systems that self-assemble into non-compact structures has proved difficult. Although several methods have been proposed, single-component nanoparticle assembly of a diamond structure has not been reported. Binary systems, in which at least one component is arranged in a diamond lattice, provide alternatives, but control of interparticle interactions is critical to this approach. DNA has been used for this purpose in a number of systems. Here we show the creation of a non-compact lattice by DNA-programmed crystallization using surface-modified Qβ phage capsid particles and gold nanoparticles, engineered to have similar effective radii. When combined with the proper connecting oligonucleotides, these components form NaTl-type colloidal crystalline structures containing interpenetrating organic and inorganic diamond lattices, as determined by small-angle X-ray scattering. DNA control of assembly is therefore shown to be compatible with particles possessing very different properties, as long as they are amenable to surface modification.

Convergence and reproducibility in molecular dynamics simulations of the DNA duplex d(GCACGAACGAACGAACGC).

PubMed

Galindo-Murillo, Rodrigo; Roe, Daniel R; Cheatham, Thomas E

2015-05-01

The structure and dynamics of DNA are critically related to its function. Molecular dynamics simulations augment experiment by providing detailed information about the atomic motions. However, to date the simulations have not been long enough for convergence of the dynamics and structural properties of DNA. Molecular dynamics simulations performed with AMBER using the ff99SB force field with the parmbsc0 modifications, including ensembles of independent simulations, were compared to long timescale molecular dynamics performed with the specialized Anton MD engine on the B-DNA structure d(GCACGAACGAACGAACGC). To assess convergence, the decay of the average RMSD values over longer and longer time intervals was evaluated in addition to assessing convergence of the dynamics via the Kullback-Leibler divergence of principal component projection histograms. These molecular dynamics simulations-including one of the longest simulations of DNA published to date at ~44μs-surprisingly suggest that the structure and dynamics of the DNA helix, neglecting the terminal base pairs, are essentially fully converged on the ~1-5μs timescale. We can now reproducibly converge the structure and dynamics of B-DNA helices, omitting the terminal base pairs, on the μs time scale with both the AMBER and CHARMM C36 nucleic acid force fields. Results from independent ensembles of simulations starting from different initial conditions, when aggregated, match the results from long timescale simulations on the specialized Anton MD engine. With access to large-scale GPU resources or the specialized MD engine "Anton" it is possible for a variety of molecular systems to reproducibly and reliably converge the conformational ensemble of sampled structures. This article is part of a Special Issue entitled: Recent developments of molecular dynamics. Copyright © 2014. Published by Elsevier B.V.

Convergence and reproducibility in molecular dynamics simulations of the DNA duplex d(GCACGAACGAACGAACGC)

PubMed Central

Galindo-Murillo, Rodrigo; Roe, Daniel R.; Cheatham, Thomas E.

2014-01-01

Background The structure and dynamics of DNA are critically related to its function. Molecular dynamics (MD) simulations augment experiment by providing detailed information about the atomic motions. However, to date the simulations have not been long enough for convergence of the dynamics and structural properties of DNA. Methods MD simulations performed with AMBER using the ff99SB force field with the parmbsc0 modifications, including ensembles of independent simulations, were compared to long timescale MD performed with the specialized Anton MD engine on the B-DNA structure d(GCACGAACGAACGAACGC). To assess convergence, the decay of the average RMSD values over longer and longer time intervals was evaluated in addition to assessing convergence of the dynamics via the Kullback-Leibler divergence of principal component projection histograms. Results These MD simulations —including one of the longest simulations of DNA published to date at ~44 μs—surprisingly suggest that the structure and dynamics of the DNA helix, neglecting the terminal base pairs, are essentially fully converged on the ~1–5 μs timescale. Conclusions We can now reproducibly converge the structure and dynamics of B-DNA helices, omitting the terminal base pairs, on the μs time scale with both the AMBER and CHARMM C36 nucleic acid force fields. Results from independent ensembles of simulations starting from different initial conditions, when aggregated, match the results from long timescale simulations on the specialized Anton MD engine. General Significance With access to large-scale GPU resources or the specialized MD engine “Anton” it is possibly for a variety of molecular systems to reproducibly and reliably converge the conformational ensemble of sampled structures. PMID:25219455

Assessing Diversity of DNA Structure-Related Sequence Features in Prokaryotic Genomes

PubMed Central

Huang, Yongjie; Mrázek, Jan

2014-01-01

Prokaryotic genomes are diverse in terms of their nucleotide and oligonucleotide composition as well as presence of various sequence features that can affect physical properties of the DNA molecule. We present a survey of local sequence patterns which have a potential to promote non-canonical DNA conformations (i.e. different from standard B-DNA double helix) and interpret the results in terms of relationships with organisms' habitats, phylogenetic classifications, and other characteristics. Our present work differs from earlier similar surveys not only by investigating a wider range of sequence patterns in a large number of genomes but also by using a more realistic null model to assess significant deviations. Our results show that simple sequence repeats and Z-DNA-promoting patterns are generally suppressed in prokaryotic genomes, whereas palindromes and inverted repeats are over-represented. Representation of patterns that promote Z-DNA and intrinsic DNA curvature increases with increasing optimal growth temperature (OGT), and decreases with increasing oxygen requirement. Additionally, representations of close direct repeats, palindromes and inverted repeats exhibit clear negative trends with increasing OGT. The observed relationships with environmental characteristics, particularly OGT, suggest possible evolutionary scenarios of structural adaptation of DNA to particular environmental niches. PMID:24408877

Mitochondrial genome-maintaining activity of mouse mitochondrial transcription factor A and its transcript isoform in Saccharomyces cerevisiae.

PubMed

Yoon, Young Geol; Koob, Michael D; Yoo, Young Hyun

2011-09-15

Mitochondrial transcription factor A (Tfam) binds to and organizes mitochondrial DNA (mtDNA) genome into a mitochondrial nucleoid (mt-nucleoid) structure, which is necessary for mtDNA transcription and maintenance. Here, we demonstrate the mtDNA-organizing activity of mouse Tfam and its transcript isoform (Tfam(iso)), which has a smaller high-mobility group (HMG)-box1 domain, using a yeast model system that contains a deletion of the yeast homolog of mouse Tfam protein, Abf2p. When the mouse Tfam genes were introduced into the ABF2 locus of yeast genome, the corresponding mouse proteins, Tfam and Tfam(iso), can functionally replace the yeast Abf2p and support mtDNA maintenance and mitochondrial biogenesis in yeast. Growth properties, mtDNA content and mitochondrial protein levels of genes encoded in the mtDNA were comparable in the strains expressing mouse proteins and the wild-type yeast strain, indicating that the proteins have robust mtDNA-maintaining and -expressing function in yeast mitochondria. These results imply that the mtDNA-organizing activities of the mouse mt-nucleoid proteins are structurally and evolutionary conserved, thus they can maintain the mtDNA of distantly related and distinctively different species, such as yeast. Copyright © 2011 Elsevier B.V. All rights reserved.

Single helically folded aromatic oligoamides that mimic the charge surface of double-stranded B-DNA

NASA Astrophysics Data System (ADS)

Ziach, Krzysztof; Chollet, Céline; Parissi, Vincent; Prabhakaran, Panchami; Marchivie, Mathieu; Corvaglia, Valentina; Bose, Partha Pratim; Laxmi-Reddy, Katta; Godde, Frédéric; Schmitter, Jean-Marie; Chaignepain, Stéphane; Pourquier, Philippe; Huc, Ivan

2018-05-01

Numerous essential biomolecular processes require the recognition of DNA surface features by proteins. Molecules mimicking these features could potentially act as decoys and interfere with pharmacologically or therapeutically relevant protein-DNA interactions. Although naturally occurring DNA-mimicking proteins have been described, synthetic tunable molecules that mimic the charge surface of double-stranded DNA are not known. Here, we report the design, synthesis and structural characterization of aromatic oligoamides that fold into single helical conformations and display a double helical array of negatively charged residues in positions that match the phosphate moieties in B-DNA. These molecules were able to inhibit several enzymes possessing non-sequence-selective DNA-binding properties, including topoisomerase 1 and HIV-1 integrase, presumably through specific foldamer-protein interactions, whereas sequence-selective enzymes were not inhibited. Such modular and synthetically accessible DNA mimics provide a versatile platform to design novel inhibitors of protein-DNA interactions.

Non-intercalative, deoxyribose binding of boric acid to calf thymus DNA.

PubMed

Ozdemir, Ayse; Gursaclı, Refiye Tekiner; Tekinay, Turgay

2014-05-01

The present study characterizes the effects of the boric acid binding on calf thymus DNA (ct-DNA) by spectroscopic and calorimetric methods. UV-Vis absorbance spectroscopy, circular dichroism (CD) spectroscopy, transmission electron microscopy (TEM), isothermal titration calorimetry (ITC), and Fourier transform infrared (FT-IR) spectroscopy were employed to characterize binding properties. Changes in the secondary structure of ct-DNA were determined by CD spectroscopy. Sizes and morphologies of boric acid-DNA complexes were determined by transmission electron microscopy (TEM). The kinetics of boric acid binding to calf thymus DNA (ct-DNA) was investigated by isothermal titration calorimetry (ITC). ITC results revealed that boric acid exhibits a moderate affinity to ct-DNA with a binding constant (K a) of 9.54 × 10(4) M(-1). FT-IR results revealed that boric acid binds to the deoxyribose sugar of DNA without disrupting the B-conformation at tested concentrations.

Synthesis, DNA Binding, and Antiproliferative Activity of Novel Acridine-Thiosemicarbazone Derivatives

PubMed Central

de Almeida, Sinara Mônica Vitalino; Lafayette, Elizabeth Almeida; Gomes da Silva, Lúcia Patrícia Bezerra; Amorim, Cézar Augusto da Cruz; de Oliveira, Tiago Bento; Gois Ruiz, Ana Lucia Tasca; de Carvalho, João Ernesto; de Moura, Ricardo Olímpio; Beltrão, Eduardo Isidoro Carneiro; de Lima, Maria do Carmo Alves; de Carvalho Júnior, Luiz Bezerra

2015-01-01

In this work, the acridine nucleus was used as a lead-compound for structural modification by adding different substituted thiosemicarbazide moieties. Eight new (Z)-2-(acridin-9-ylmethylene)-N-phenylhydrazinecarbothioamide derivatives (3a–h) were synthesized, their antiproliferative activities were evaluated, and DNA binding properties were performed with calf thymus DNA (ctDNA) by electronic absorption and fluorescence spectroscopies. Both hyperchromic and hypochromic effects, as well as red or blue shifts were demonstrated by addition of ctDNA to the derivatives. The calculated binding constants ranged from 1.74 × 104 to 1.0 × 106 M−1 and quenching constants from −0.2 × 104 to 2.18 × 104 M−1 indicating high affinity to ctDNA base pairs. The most efficient compound in binding to ctDNA in vitro was (Z)-2-(acridin-9-ylmethylene)-N-(4-chlorophenyl) hydrazinecarbothioamide (3f), while the most active compound in antiproliferative assay was (Z)-2-(acridin-9-ylmethylene)-N-phenylhydrazinecarbothioamide (3a). There was no correlation between DNA-binding and in vitro antiproliferative activity, but the results suggest that DNA binding can be involved in the biological activity mechanism. This study may guide the choice of the size and shape of the intercalating part of the ligand and the strategic selection of substituents that increase DNA-binding or antiproliferative properties. PMID:26068233

Synthesis and dielectric properties of zinc oxide nanoparticles using a biotemplate

DOE Office of Scientific and Technical Information (OSTI.GOV)

P, Sharmila P, E-mail: sharmilavishram@gmail.com; Tharayil, Nisha J., E-mail: nishajohntharayil@gmail.com

Zinc Oxide nanoparticles are synthesized using DNA as capping agent. Zinc oxide nanoparticles are synthesized using DNA as a capping agent. Structural and morphological characterizations are done using SEM, FTIR and XRD. The particle size and lattice parameters are calculated from the diffraction data. The optical properties are studied using UV-Vis absorption spectroscopy and bandgap variation with temperature is determined. The dielectric property of nanoparticles is studied by varying temperature and frequency. The dielectric constant and dispersion parameters are found out. Method of Cole-Cole analysis is used to study the high temperature dispersion of relaxation time. The variation of bothmore » AC and DC conductivity are studied and activation energy calculated.« less

Comparing the Properties of Electrochemical-Based DNA Sensors Employing Different Redox Tags

PubMed Central

Kang, Di; Zuo, Xiaolei; Yang, Renqiang; Xia, Fan; Plaxco, Kevin W.; White, Ryan J.

2009-01-01

Many electrochemical biosensor approaches developed in recent years utilize redox labeled (most commonly methylene blue or ferrocene) oligonucleotide probes site-specifically attached to an interrogating electrode. Sensors in this class have been reported employing a range of probe architectures, including single- and double-stranded DNA, more complex DNA structures, DNA and RNA aptamers and, most recently, DNA-small molecule chimeras. Signaling in this class of sensors is generally predicated on binding-induced changes in the efficiency with which the covalently attached redox label transfers electrons with the interrogating electrode. Here we have investigated how the properties of the redox tag affect the performance of such sensors. Specifically, we compare the differences in signaling and stability of electrochemical DNA sensors (E-DNA sensors) fabricated using either ferrocene or methylene blue as the signaling redox moiety. We find that while both tags support efficient E-DNA signaling, ferrocene produces slightly improved signal gain and target affinity. These small advantages, however, come at a potentially significant price: the ferrocene-based sensors are far less stable than their methylene blue counterparts, particularly with regards to stability to long-term storage, repeated electrochemical interrogations, repeated sensing/regeneration iterations, and employment in complex sample matrices such as blood serum. PMID:19810694

Self-assembly of nucleic acids, silk and hybrid materials thereof.

PubMed

Humenik, Martin; Scheibel, Thomas

2014-12-17

Top-down approaches based on etching techniques have almost reached their limits in terms of dimension. Therefore, novel assembly strategies and types of nanomaterials are required to allow technological advances. Self-assembly processes independent of external energy sources and unlimited in dimensional scaling have become a very promising approach. Here,we highlight recent developments in self-assembled DNA-polymer, silk-polymer and silk-DNA hybrids as promising materials with biotic and abiotic moieties for constructing complex hierarchical materials in ‘bottom-up’ approaches. DNA block copolymers assemble into nanostructures typically exposing a DNA corona which allows functionalization, labeling and higher levels of organization due to its specific addressable recognition properties. In contrast, self-assembly of natural silk proteins as well as their recombinant variants yields mechanically stable β-sheet rich nanostructures. The combination of silk with abiotic polymers gains hybrid materials with new functionalities. Together, the precision of DNA hybridization and robustness of silk fibrillar structures combine in novel conjugates enable processing of higher-order structures with nanoscale architecture and programmable functions.

Self-assembly of nucleic acids, silk and hybrid materials thereof

NASA Astrophysics Data System (ADS)

Humenik, Martin; Scheibel, Thomas

2014-12-01

Top-down approaches based on etching techniques have almost reached their limits in terms of dimension. Therefore, novel assembly strategies and types of nanomaterials are required to allow technological advances. Self-assembly processes independent of external energy sources and unlimited in dimensional scaling have become a very promising approach. Here, we highlight recent developments in self-assembled DNA-polymer, silk-polymer and silk-DNA hybrids as promising materials with biotic and abiotic moieties for constructing complex hierarchical materials in ‘bottom-up’ approaches. DNA block copolymers assemble into nanostructures typically exposing a DNA corona which allows functionalization, labeling and higher levels of organization due to its specific addressable recognition properties. In contrast, self-assembly of natural silk proteins as well as their recombinant variants yields mechanically stable β-sheet rich nanostructures. The combination of silk with abiotic polymers gains hybrid materials with new functionalities. Together, the precision of DNA hybridization and robustness of silk fibrillar structures combine in novel conjugates enable processing of higher-order structures with nanoscale architecture and programmable functions.

Interaction of cationic phthalocyanines with DNA. Importance of the structure of the substituents.

PubMed

López Zeballos, N C; Gauna, G A; García Vior, M C; Awruch, J; Dicelio, L E

2014-07-05

The interaction of novel zinc (II) cationic phthalocyanines with CT-DNA was studied using absorption and fluorescence spectroscopy, as well as thermal denaturation profiles. Results showed an electrostatic interaction between the phthalocyanines and CT-DNA. The properties of these phthalocyanines were compared taking the structure of the macrocycle peripheral substituents into account. 2,9(10),16(17),23(24)-tetrakis[(N-butyl-N-methylammonium)ethylsulfanyl]phthalocyaninatozinc(II) tetraiodide (Pc6) had a greater affinity for the CT-DNA helix than its bioisoster 2,9(10),16(17),23(24)-tetrakis[(N-dibutyl-N-methylammonium)ethoxy]phthalocyaninatozinc(II) tetraiodide (Pc7). 2,9(10),16(17),23(24)-tetrakis[(2-trimethylammonium)ethyl-sulfanyl]phthalocyaninatozinc(II) tetraiodide (Pc13) also carried a sulfur atom like Pc6, but linked to bulky substituents such as trimethylammonium groups. The planar aromatic region of the cationic phthalocyanines in this study appears to be unable to facilitate their intercalation with CT-DNA. Copyright © 2014 Elsevier B.V. All rights reserved.

Nanomechanical molecular devices made of DNA origami.

PubMed

Kuzuya, Akinori; Ohya, Yuichi

2014-06-17

CONSPECTUS: Eight years have passed since the striking debut of the DNA origami technique ( Rothemund, P. W. K. Nature 2006 , 440 , 297 - 302 ), in which long single-stranded DNA is folded into a designed nanostructure, in either 2D or 3D, with the aid of many short staple strands. The number of proposals for new design principles for DNA origami structures seems to have already reached a peak. It is apparent that DNA origami study is now entering the second phase of creating practical applications. The development of functional nanomechanical molecular devices using the DNA origami technique is one such application attracting significant interest from researchers in the field. Nanomechanical DNA origami devices, which maintain the characteristics of DNA origami structures, have various advantages over conventional DNA nanomachines. Comparatively high assembly yield, relatively large size visible via atomic force microscopy (AFM) or transmission electron microscopy (TEM), and the capability to assemble multiple functional groups with precision using multiple staple strands are some of the advantages of the DNA origami technique for constructing sophisticated molecular devices. This Account describes the recent developments of such nanomechanical DNA origami devices and reviews the emerging target of DNA origami studies. First, simple "dynamic" DNA origami structures with transformation capability, such as DNA origami boxes and a DNA origami hatch with structure control, are briefly summarized. More elaborate nanomechanical DNA origami devices are then reviewed. The first example describes DNA origami pinching devices that can be used as "single-molecule" beacons to detect a variety of biorelated molecules, from metal ions at the size of a few tens of atomic mass number units to relatively gigantic proteins with a molecular mass greater than a hundred kilodaltons, all on a single platform. Clamshell-like DNA nanorobots equipped with logic gates can discriminate different cell lines, open their shell, and bind to their target. An intelligent DNA origami "sheath" can mimic the function of suppressors in a transcription regulation system to control the expression of a loaded gene. DNA origami "rolls" are created to construct precisely arranged plasmonic devices with metal nanoparticles. All of their functions are derived from their nanomechanical movement, which is programmable by designing the DNA sequence or by using the significant repository of technical achievements in nucleic acid chemistry. Finally, some studies on detailed structural parameters of DNA origami or their mechanical properties in nanoscale are discussed, which may be useful and inspiring for readers who intend to design new nanomechanical DNA origami devices.

Transposon-like properties of the major, long repetitive sequence family in the genome of Physarum polycephalum

PubMed Central

Pearston, Douglas H.; Gordon, Mairi; Hardman, Norman

1985-01-01

A family of long, highly-repetitive sequences, referred to previously as `HpaII-repeats', dominates the genome of the eukaryotic slime mould Physarum polycephalum. These sequences are found exclusively in scrambled clusters. They account for about one-half of the total complement of repetitive DNA in Physarum, and represent the major sequence component found in hypermethylated, 20-50 kb segments of Physarum genomic DNA that fail to be cleaved using the restriction endonuclease HpaII. The structure of this abundant repetitive element was investigated by analysing cloned segments derived from the hypermethylated genomic DNA compartment. We show that the `HpaII-repeat' forms part of a larger repetitive DNA structure, ∼8.6 kb in length, with several structural features in common with recognised eukaryotic transposable genetic elements. Scrambled clusters of the sequence probably arise as a result of transposition-like events, during which the element preferentially recombines in either orientation with target sites located in other copies of the same repeated sequence. The target sites for transposition/recombination are not related in sequence but in all cases studied they are potentially capable of promoting the formation of small `cruciforms' or `Z-DNA' structures which might be recognised during the recombination process. ImagesFig. 3.Fig. 4. PMID:16453652

Detection of Ammonia-Oxidizing Bacteria (AOB) Using a Porous Silicon Optical Biosensor Based on a Multilayered Double Bragg Mirror Structure.

PubMed

Zhang, Hongyan; Lv, Jie; Jia, Zhenhong

2018-01-01

We successfully demonstrate a porous silicon (PS) double Bragg mirror by electrochemical etching at room temperature as a deoxyribonucleic acid (DNA) label-free biosensor for detecting ammonia-oxidizing bacteria (AOB). Compared to various other one-dimension photonic crystal configurations of PS, the double Bragg mirror structure is quite easy to prepare and exhibits interesting optical properties. The width of high reflectivity stop band of the PS double Bragg mirror is about 761 nm with a sharp and deep resonance peak at 1328 nm in the reflectance spectrum, which gives a high sensitivity and distinguishability for sensing performance. The detection sensitivity of such a double Bragg mirror structure is illustrated through the investigation of AOB DNA hybridization in the PS pores. The redshifts of the reflectance spectra show a good linear relationship with both complete complementary and partial complementary DNA. The lowest detection limit for complete complementary DNA is 27.1 nM and the detection limit of the biosensor for partial complementary DNA is 35.0 nM, which provides the feasibility and effectiveness for the detection of AOB in a real environment. The PS double Bragg mirror structure is attractive for widespread biosensing applications and provides great potential for the development of optical applications.

Natural Compounds as Anticancer Agents Targeting DNA Topoisomerases

PubMed Central

Jain, Chetan Kumar; Majumder, Hemanta Kumar; Roychoudhury, Susanta

2017-01-01

DNA topoisomerases are important cellular enzymes found in almost all types of living cells (eukaryotic and prokaryotic). These enzymes are essential for various DNA metabolic processes e.g. replication, transcription, recombination, chromosomal decatenation etc. These enzymes are important molecular drug targets and inhibitors of these enzymes are widely used as effective anticancer and antibacterial drugs. However, topoisomerase inhibitors have some therapeutic limitations and they exert serious side effects during cancer chemotherapy. Thus, development of novel anticancer topoisomerase inhibitors is necessary for improving cancer chemotherapy. Nature serves as a repertoire of structurally and chemically diverse molecules and in the recent years many DNA topoisomerase inhibitors have been identified from natural sources. The present review discusses anticancer properties and therapeutic importance of eighteen recently identified natural topoisomerase inhibitors (from the year 2009 to 2015). Structural characteristics of these novel inhibitors provide backbones for designing and developing new anticancer drugs. PMID:28503091

Mosaic organization of DNA nucleotides

NASA Technical Reports Server (NTRS)

Peng, C. K.; Buldyrev, S. V.; Havlin, S.; Simons, M.; Stanley, H. E.; Goldberger, A. L.

1994-01-01

Long-range power-law correlations have been reported recently for DNA sequences containing noncoding regions. We address the question of whether such correlations may be a trivial consequence of the known mosaic structure ("patchiness") of DNA. We analyze two classes of controls consisting of patchy nucleotide sequences generated by different algorithms--one without and one with long-range power-law correlations. Although both types of sequences are highly heterogenous, they are quantitatively distinguishable by an alternative fluctuation analysis method that differentiates local patchiness from long-range correlations. Application of this analysis to selected DNA sequences demonstrates that patchiness is not sufficient to account for long-range correlation properties.

Nanoarchitectonics with Porphyrin Functionalized DNA

PubMed Central

2017-01-01

Conspectus DNA is well-known as bearer of the genetic code. Since its structure elucidation nearly seven decades ago by Watson, Crick, Wilkins, and Franklin, much has been learned about its detailed structure, function, and genetic coding. The development of automated solid-phase synthesis, and with it the availability of synthetic DNA with any desired sequence in lengths of up to hundreds of bases in the best case, has contributed much to the advancement of the field of DNA research. In addition, classic organic synthesis has allowed introduction of a very large number of modifications in the DNA in a sequence specific manner, which have initially been targeted at altering the biological function of DNA. However, in recent years DNA has become a very attractive scaffold in supramolecular chemistry, where DNA is taken out of its biological role and serves as both stick and glue molecule to assemble novel functional structures with nanometer precision. The attachment of functionalities to DNA has led to the creation of supramolecular systems with applications in light harvesting, energy and electron transfer, sensing, and catalysis. Functional DNA is clearly having a significant impact in the field of bioinspired nanosystems. Of particular interest is the use of porphyrins in supramolecular chemistry and bionanotechnology, because they are excellent functional groups due to their electronic properties that can be tailored through chemical modifications of the aromatic core or through insertion of almost any metal of the periodic table into the central cavity. The porphyrins can be attached either to the nucleobase, to the phosphate group, or to the ribose moiety. Additionally, noncovalent templating through Watson–Crick base pairing forms an alternative and attractive approach. With this, the combination of two seemingly simple molecules gives rise to a highly complex system with unprecedented possibilities for modulation of function, and with it applications, particularly when combined with other functional groups. Here, an overview is given on the developments of using porphyrin modified DNA for the construction of functional assemblies. Strategies for the synthesis and characterization are presented alongside selected applications where the porphyrin modification has proven to be particularly useful and superior to other modifiers but also has revealed its limitations. We also discuss implications on properties and behavior of the porphyrin–DNA, where similar issues could arise when using other hydrophobic and bulky substituents on DNA. This includes particularly problems regarding synthesis of the building blocks, DNA synthesis, yields, solubility, and intermolecular interactions. PMID:28272871

DNA nanotubes for NMR structure determination of membrane proteins.

PubMed

Bellot, Gaëtan; McClintock, Mark A; Chou, James J; Shih, William M

2013-04-01

Finding a way to determine the structures of integral membrane proteins using solution nuclear magnetic resonance (NMR) spectroscopy has proved to be challenging. A residual-dipolar-coupling-based refinement approach can be used to resolve the structure of membrane proteins up to 40 kDa in size, but to do this you need a weak-alignment medium that is detergent-resistant and it has thus far been difficult to obtain such a medium suitable for weak alignment of membrane proteins. We describe here a protocol for robust, large-scale synthesis of detergent-resistant DNA nanotubes that can be assembled into dilute liquid crystals for application as weak-alignment media in solution NMR structure determination of membrane proteins in detergent micelles. The DNA nanotubes are heterodimers of 400-nm-long six-helix bundles, each self-assembled from a M13-based p7308 scaffold strand and >170 short oligonucleotide staple strands. Compatibility with proteins bearing considerable positive charge as well as modulation of molecular alignment, toward collection of linearly independent restraints, can be introduced by reducing the negative charge of DNA nanotubes using counter ions and small DNA-binding molecules. This detergent-resistant liquid-crystal medium offers a number of properties conducive for membrane protein alignment, including high-yield production, thermal stability, buffer compatibility and structural programmability. Production of sufficient nanotubes for four or five NMR experiments can be completed in 1 week by a single individual.

A Case Study into Microbial Genome Assembly Gap Sequences and Finishing Strategies.

PubMed

Utturkar, Sagar M; Klingeman, Dawn M; Hurt, Richard A; Brown, Steven D

2017-01-01

This study characterized regions of DNA which remained unassembled by either PacBio and Illumina sequencing technologies for seven bacterial genomes. Two genomes were manually finished using bioinformatics and PCR/Sanger sequencing approaches and regions not assembled by automated software were analyzed. Gaps present within Illumina assemblies mostly correspond to repetitive DNA regions such as multiple rRNA operon sequences. PacBio gap sequences were evaluated for several properties such as GC content, read coverage, gap length, ability to form strong secondary structures, and corresponding annotations. Our hypothesis that strong secondary DNA structures blocked DNA polymerases and contributed to gap sequences was not accepted. PacBio assemblies had few limitations overall and gaps were explained as cumulative effect of lower than average sequence coverage and repetitive sequences at contig termini. An important aspect of the present study is the compilation of biological features that interfered with assembly and included active transposons, multiple plasmid sequences, phage DNA integration, and large sequence duplication. Our targeted genome finishing approach and systematic evaluation of the unassembled DNA will be useful for others looking to close, finish, and polish microbial genome sequences.

iSS-PseDNC: identifying splicing sites using pseudo dinucleotide composition.

PubMed

Chen, Wei; Feng, Peng-Mian; Lin, Hao; Chou, Kuo-Chen

2014-01-01

In eukaryotic genes, exons are generally interrupted by introns. Accurately removing introns and joining exons together are essential processes in eukaryotic gene expression. With the avalanche of genome sequences generated in the postgenomic age, it is highly desired to develop automated methods for rapid and effective detection of splice sites that play important roles in gene structure annotation and even in RNA splicing. Although a series of computational methods were proposed for splice site identification, most of them neglected the intrinsic local structural properties. In the present study, a predictor called "iSS-PseDNC" was developed for identifying splice sites. In the new predictor, the sequences were formulated by a novel feature-vector called "pseudo dinucleotide composition" (PseDNC) into which six DNA local structural properties were incorporated. It was observed by the rigorous cross-validation tests on two benchmark datasets that the overall success rates achieved by iSS-PseDNC in identifying splice donor site and splice acceptor site were 85.45% and 87.73%, respectively. It is anticipated that iSS-PseDNC may become a useful tool for identifying splice sites and that the six DNA local structural properties described in this paper may provide novel insights for in-depth investigations into the mechanism of RNA splicing.

DNA Polymorphism: A Comparison of Force Fields for Nucleic Acids

PubMed Central

Reddy, Swarnalatha Y.; Leclerc, Fabrice; Karplus, Martin

2003-01-01

The improvements of the force fields and the more accurate treatment of long-range interactions are providing more reliable molecular dynamics simulations of nucleic acids. The abilities of certain nucleic acid force fields to represent the structural and conformational properties of nucleic acids in solution are compared. The force fields are AMBER 4.1, BMS, CHARMM22, and CHARMM27; the comparison of the latter two is the primary focus of this paper. The performance of each force field is evaluated first on its ability to reproduce the B-DNA decamer d(CGATTAATCG)2 in solution with simulations in which the long-range electrostatics were treated by the particle mesh Ewald method; the crystal structure determined by Quintana et al. (1992) is used as the starting point for all simulations. A detailed analysis of the structural and solvation properties shows how well the different force fields can reproduce sequence-specific features. The results are compared with data from experimental and previous theoretical studies. PMID:12609851

Distinct epigenomic landscapes of pluripotent and lineage-committed human cells.

PubMed

Hawkins, R David; Hon, Gary C; Lee, Leonard K; Ngo, Queminh; Lister, Ryan; Pelizzola, Mattia; Edsall, Lee E; Kuan, Samantha; Luu, Ying; Klugman, Sarit; Antosiewicz-Bourget, Jessica; Ye, Zhen; Espinoza, Celso; Agarwahl, Saurabh; Shen, Li; Ruotti, Victor; Wang, Wei; Stewart, Ron; Thomson, James A; Ecker, Joseph R; Ren, Bing

2010-05-07

Human embryonic stem cells (hESCs) share an identical genome with lineage-committed cells, yet possess the remarkable properties of self-renewal and pluripotency. The diverse cellular properties in different cells have been attributed to their distinct epigenomes, but how much epigenomes differ remains unclear. Here, we report that epigenomic landscapes in hESCs and lineage-committed cells are drastically different. By comparing the chromatin-modification profiles and DNA methylomes in hESCs and primary fibroblasts, we find that nearly one-third of the genome differs in chromatin structure. Most changes arise from dramatic redistributions of repressive H3K9me3 and H3K27me3 marks, which form blocks that significantly expand in fibroblasts. A large number of potential regulatory sequences also exhibit a high degree of dynamics in chromatin modifications and DNA methylation. Additionally, we observe novel, context-dependent relationships between DNA methylation and chromatin modifications. Our results provide new insights into epigenetic mechanisms underlying properties of pluripotency and cell fate commitment.

Quantum Mechanical Modeling of Ballistic MOSFETs

NASA Technical Reports Server (NTRS)

Svizhenko, Alexei; Anantram, M. P.; Govindan, T. R.; Biegel, Bryan (Technical Monitor)

2001-01-01

The objective of this project was to develop theory, approximations, and computer code to model quasi 1D structures such as nanotubes, DNA, and MOSFETs: (1) Nanotubes: Influence of defects on ballistic transport, electro-mechanical properties, and metal-nanotube coupling; (2) DNA: Model electron transfer (biochemistry) and transport experiments, and sequence dependence of conductance; and (3) MOSFETs: 2D doping profiles, polysilicon depletion, source to drain and gate tunneling, understand ballistic limit.

Density functional study on the structural and thermodynamic properties of aqueous DNA-electrolyte solution in the framework of cell model.

PubMed

Wang, Ke; Yu, Yang-Xin; Gao, Guang-Hua

2008-05-14

A density functional theory (DFT) in the framework of cell model is proposed to calculate the structural and thermodynamic properties of aqueous DNA-electrolyte solution with finite DNA concentrations. The hard-sphere contribution to the excess Helmholtz energy functional is derived from the modified fundamental measure theory, and the electrostatic interaction is evaluated through a quadratic functional Taylor expansion around a uniform fluid. The electroneutrality in the cell leads to a variational equation with a constraint. Since the reference fluid is selected to be a bulk phase, the Lagrange multiplier proves to be the potential drop across the cell boundary (Donnan potential). The ion profiles and electrostatic potential profiles in the cell are calculated from the present DFT-cell model. Our DFT-cell model gives better prediction of ion profiles than the Poisson-Boltzmann (PB)- or modified PB-cell models when compared to the molecular simulation data. The effects of polyelectrolyte concentration, ion size, and added-salt concentration on the electrostatic potential difference between the DNA surface and the cell boundary are investigated. The expression of osmotic coefficient is derived from the general formula of grand potential. The osmotic coefficients predicted by the DFT are lower than the PB results and are closer to the simulation results and experimental data.

Interaction of vasicine with calf thymus DNA: Molecular docking, spectroscopic and differential scanning calorimetric insights.

PubMed

R S, Sai Murali; R S, Sai Siddhardha; D, Rajesh Babu; S, Venketesh; R, Basavaraju; G, Nageswara Rao

2017-06-05

The present study brings out the interaction between vasicine, an alkaloid and Adhatoda vasica Nees with double stranded DNA. The physico-chemical interaction between small molecules and nucleic acids is a major area of focus in screening drugs against various cancers. Molecular probing in our study using Molecular Operating Environment (MOE) has revealed interaction of vasicine with DNA double helix. Here we report the interaction of vasicine with Calf thymus DNA. We present for the first time the results obtained from UV-visible, fluorescence spectroscopic and differential scanning calorimetric techniques that suggest a moderate to strong electrostatic, hydrophobic and van der Waals interactions mediating the DNA binding properties of vasicine, leading to disruption of DNA secondary structure. Copyright © 2017 Elsevier B.V. All rights reserved.

Identification of Specific DNA Binding Residues in the TCP Family of Transcription Factors in Arabidopsis[W

PubMed Central

Aggarwal, Pooja; Das Gupta, Mainak; Joseph, Agnel Praveen; Chatterjee, Nirmalya; Srinivasan, N.; Nath, Utpal

2010-01-01

The TCP transcription factors control multiple developmental traits in diverse plant species. Members of this family share an ∼60-residue-long TCP domain that binds to DNA. The TCP domain is predicted to form a basic helix-loop-helix (bHLH) structure but shares little sequence similarity with canonical bHLH domain. This classifies the TCP domain as a novel class of DNA binding domain specific to the plant kingdom. Little is known about how the TCP domain interacts with its target DNA. We report biochemical characterization and DNA binding properties of a TCP member in Arabidopsis thaliana, TCP4. We have shown that the 58-residue domain of TCP4 is essential and sufficient for binding to DNA and possesses DNA binding parameters comparable to canonical bHLH proteins. Using a yeast-based random mutagenesis screen and site-directed mutants, we identified the residues important for DNA binding and dimer formation. Mutants defective in binding and dimerization failed to rescue the phenotype of an Arabidopsis line lacking the endogenous TCP4 activity. By combining structure prediction, functional characterization of the mutants, and molecular modeling, we suggest a possible DNA binding mechanism for this class of transcription factors. PMID:20363772

Novel structural features drive DNA binding properties of Cmr, a CRP family protein in TB complex mycobacteria.

PubMed

Ranganathan, Sridevi; Cheung, Jonah; Cassidy, Michael; Ginter, Christopher; Pata, Janice D; McDonough, Kathleen A

2018-01-09

Mycobacterium tuberculosis (Mtb) encodes two CRP/FNR family transcription factors (TF) that contribute to virulence, Cmr (Rv1675c) and CRPMt (Rv3676). Prior studies identified distinct chromosomal binding profiles for each TF despite their recognizing overlapping DNA motifs. The present study shows that Cmr binding specificity is determined by discriminator nucleotides at motif positions 4 and 13. X-ray crystallography and targeted mutational analyses identified an arginine-rich loop that expands Cmr's DNA interactions beyond the classical helix-turn-helix contacts common to all CRP/FNR family members and facilitates binding to imperfect DNA sequences. Cmr binding to DNA results in a pronounced asymmetric bending of the DNA and its high level of cooperativity is consistent with DNA-facilitated dimerization. A unique N-terminal extension inserts between the DNA binding and dimerization domains, partially occluding the site where the canonical cAMP binding pocket is found. However, an unstructured region of this N-terminus may help modulate Cmr activity in response to cellular signals. Cmr's multiple levels of DNA interaction likely enhance its ability to integrate diverse gene regulatory signals, while its novel structural features establish Cmr as an atypical CRP/FNR family member. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Studies of torsional properties of DNA and nucleosomes using angular optical trapping

NASA Astrophysics Data System (ADS)

Sheinin, Maxim Y.

DNA in vivo is subjected to torsional stress due to the action of molecular motors and other DNA-binding proteins. Several decades of research have uncovered the fascinating diversity of DNA transformations under torsion and the important role they play in the regulation of vital cellular processes such as transcription and replication. Recent studies have also suggested that torsion can influence the structure and stability of nucleosomes---basic building blocks of the eukaryotic genome. However, our understanding of the impact of torsion is far from being complete due to significant experimental challenges. In this work we have used a powerful single-molecule experimental technique, angular optical trapping, to address several long-standing issues in the field of DNA and nucleosome mechanics. First, we utilized the high resolution and direct torque measuring capability of the angular optical trapping to precisely measure DNA twist-stretch coupling. Second, we characterized DNA melting under tension and torsion. We found that torsionally underwound DNA forms a left-handed structure, significantly more flexible compared to the regular B-DNA. Finally, we performed the first comprehensive investigation of the single nucleosome behavior under torque and force. Importantly, we discovered that positive torque causes significant dimer loss, which can have implications for transcription through chromatin.

The Role of Structural Enthalpy in Spherical Nucleic Acid Hybridization.

PubMed

Fong, Lam-Kiu; Wang, Ziwei; Schatz, George C; Luijten, Erik; Mirkin, Chad A

2018-05-23

DNA hybridization onto DNA-functionalized nanoparticle surfaces (e.g., in the form of a spherical nucleic acid (SNA)) is known to be enhanced relative to hybridization free in solution. Surprisingly, via isothermal titration calorimetry, we reveal that this enhancement is enthalpically, as opposed to entropically, dominated by ∼20 kcal/mol. Coarse-grained molecular dynamics simulations suggest that the observed enthalpic enhancement results from structurally confining the DNA on the nanoparticle surface and preventing it from adopting enthalpically unfavorable conformations like those observed in the solution case. The idea that structural confinement leads to the formation of energetically more stable duplexes is evaluated by decreasing the degree of confinement a duplex experiences on the nanoparticle surface. Both experiment and simulation confirm that when the surface-bound duplex is less confined, i.e., at lower DNA surface density or at greater distance from the nanoparticle surface, its enthalpy of formation approaches the less favorable enthalpy of duplex formation for the linear strand in solution. This work provides insight into one of the most important and enabling properties of SNAs and will inform the design of materials that rely on the thermodynamics of hybridization onto DNA-functionalized surfaces, including diagnostic probes and therapeutic agents.

In vitro fluorescence studies of transcription factor IIB-DNA interaction.

PubMed

Górecki, Andrzej; Figiel, Małgorzata; Dziedzicka-Wasylewska, Marta

2015-01-01

General transcription factor TFIIB is one of the basal constituents of the preinitiation complex of eukaryotic RNA polymerase II, acting as a bridge between the preinitiation complex and the polymerase, and binding promoter DNA in an asymmetric manner, thereby defining the direction of the transcription. Methods of fluorescence spectroscopy together with circular dichroism spectroscopy were used to observe conformational changes in the structure of recombinant human TFIIB after binding to specific DNA sequence. To facilitate the exploration of the structural changes, several site-directed mutations have been introduced altering the fluorescence properties of the protein. Our observations showed that binding of specific DNA sequences changed the protein structure and dynamics, and TFIIB may exist in two conformational states, which can be described by a different microenvironment of W52. Fluorescence studies using both intrinsic and exogenous fluorophores showed that these changes significantly depended on the recognition sequence and concerned various regions of the protein, including those interacting with other transcription factors and RNA polymerase II. DNA binding can cause rearrangements in regions of proteins interacting with the polymerase in a manner dependent on the recognized sequences, and therefore, influence the gene expression.

Structure and electrical properties of DNA nanotubes embedded in lipid bilayer membranes

PubMed Central

Maiti, Prabal K

2018-01-01

Abstract Engineering the synthetic nanopores through lipid bilayer membrane to access the interior of a cell is a long persisting challenge in biotechnology. Here, we demonstrate the stability and dynamics of a tile-based 6-helix DNA nanotube (DNT) embedded in POPC lipid bilayer using the analysis of 0.2 μs long equilibrium MD simulation trajectories. We observe that the head groups of the lipid molecules close to the lumen cooperatively tilt towards the hydrophilic sugar-phosphate backbone of DNA and form a toroidal structure around the patch of DNT protruding in the membrane. Further, we explore the effect of ionic concentrations to the in-solution structure and stability of the lipid-DNT complex. Transmembrane ionic current measurements for the constant electric field MD simulation provide the I-V characteristics of the water filled DNT lumen in lipid membrane. With increasing salt concentrations, the measured values of transmembrane ionic conductance of the porous DNT lumen vary from 4.3 to 20.6 nS. Simulations of the DNTs with ssDNA and dsDNA overhangs at the mouth of the pore show gating effect with remarkable difference in the transmembrane ionic conductivities for open and close state nanopores. PMID:29136243

The intervening domain from MeCP2 enhances the DNA affinity of the methyl binding domain and provides an independent DNA interaction site.

PubMed

Claveria-Gimeno, Rafael; Lanuza, Pilar M; Morales-Chueca, Ignacio; Jorge-Torres, Olga C; Vega, Sonia; Abian, Olga; Esteller, Manel; Velazquez-Campoy, Adrian

2017-01-31

Methyl-CpG binding protein 2 (MeCP2) preferentially interacts with methylated DNA and it is involved in epigenetic regulation and chromatin remodelling. Mutations in MeCP2 are linked to Rett syndrome, the leading cause of intellectual retardation in girls and causing mental, motor and growth impairment. Unstructured regions in MeCP2 provide the plasticity for establishing interactions with multiple binding partners. We present a biophysical characterization of the methyl binding domain (MBD) from MeCP2 reporting the contribution of flanking domains to its structural stability and dsDNA interaction. The flanking disordered intervening domain (ID) increased the structural stability of MBD, modified its dsDNA binding profile from an entropically-driven moderate-affinity binding to an overwhelmingly enthalpically-driven high-affinity binding. Additionally, ID provided an additional site for simultaneously and autonomously binding an independent dsDNA molecule, which is a key feature linked to the chromatin remodelling and looping activity of MeCP2, as well as its ability to interact with nucleosomes replacing histone H1. The dsDNA interaction is characterized by an unusually large heat capacity linked to a cluster of water molecules trapped within the binding interface. The dynamics of disordered regions together with extrinsic factors are key determinants of MeCP2 global structural properties and functional capabilities.

The intervening domain from MeCP2 enhances the DNA affinity of the methyl binding domain and provides an independent DNA interaction site

PubMed Central

Claveria-Gimeno, Rafael; Lanuza, Pilar M.; Morales-Chueca, Ignacio; Jorge-Torres, Olga C.; Vega, Sonia; Abian, Olga; Esteller, Manel; Velazquez-Campoy, Adrian

2017-01-01

Methyl-CpG binding protein 2 (MeCP2) preferentially interacts with methylated DNA and it is involved in epigenetic regulation and chromatin remodelling. Mutations in MeCP2 are linked to Rett syndrome, the leading cause of intellectual retardation in girls and causing mental, motor and growth impairment. Unstructured regions in MeCP2 provide the plasticity for establishing interactions with multiple binding partners. We present a biophysical characterization of the methyl binding domain (MBD) from MeCP2 reporting the contribution of flanking domains to its structural stability and dsDNA interaction. The flanking disordered intervening domain (ID) increased the structural stability of MBD, modified its dsDNA binding profile from an entropically-driven moderate-affinity binding to an overwhelmingly enthalpically-driven high-affinity binding. Additionally, ID provided an additional site for simultaneously and autonomously binding an independent dsDNA molecule, which is a key feature linked to the chromatin remodelling and looping activity of MeCP2, as well as its ability to interact with nucleosomes replacing histone H1. The dsDNA interaction is characterized by an unusually large heat capacity linked to a cluster of water molecules trapped within the binding interface. The dynamics of disordered regions together with extrinsic factors are key determinants of MeCP2 global structural properties and functional capabilities. PMID:28139759

Fluctuations in the DNA double helix

NASA Astrophysics Data System (ADS)

Peyrard, M.; López, S. C.; Angelov, D.

2007-08-01

DNA is not the static entity suggested by the famous double helix structure. It shows large fluctuational openings, in which the bases, which contain the genetic code, are temporarily open. Therefore it is an interesting system to study the effect of nonlinearity on the physical properties of a system. A simple model for DNA, at a mesoscopic scale, can be investigated by computer simulation, in the same spirit as the original work of Fermi, Pasta and Ulam. These calculations raise fundamental questions in statistical physics because they show a temporary breaking of equipartition of energy, regions with large amplitude fluctuations being able to coexist with regions where the fluctuations are very small, even when the model is studied in the canonical ensemble. This phenomenon can be related to nonlinear excitations in the model. The ability of the model to describe the actual properties of DNA is discussed by comparing theoretical and experimental results for the probability that base pairs open an a given temperature in specific DNA sequences. These studies give us indications on the proper description of the effect of the sequence in the mesoscopic model.

DNA interactions of non-chelating tinidazole-based coordination compounds and their structural, redox and cytotoxic properties.

PubMed

Castro-Ramírez, Rodrigo; Ortiz-Pastrana, Naytzé; Caballero, Ana B; Zimmerman, Matthew T; Stadelman, Bradley S; Gaertner, Andrea A E; Brumaghim, Julia L; Korrodi-Gregório, Luís; Pérez-Tomás, Ricardo; Gamez, Patrick; Barba-Behrens, Norah

2018-05-23

Novel tinidazole (tnz) coordination compounds of different geometries were synthesised, whose respective solid-state packing appears to be driven by inter- and intramolecular lone pairπ interactions. The copper(ii) compounds exhibit interesting redox properties originating from both the tnz and the metal ions. These complexes interact with DNA through two distinct ways, namely via electrostatic interactions or/and groove binding, and they can mediate the generation of ROS that damage the biomolecule. Cytotoxic studies revealed an interesting activity of the dinuclear compound [Cu(tnz)2(μ-Cl)Cl]2 7, which is further more efficient towards cancer cells, compared with normal cells.

The NnCenH3 protein and centromeric DNA sequence profiles of Nelumbo nucifera Gaertn. (sacred lotus) reveal the DNA structures and dynamics of centromeres in basal eudicots.

PubMed

Zhu, Zhixuan; Gui, Songtao; Jin, Jing; Yi, Rong; Wu, Zhihua; Qian, Qian; Ding, Yi

2016-09-01

Centromeres on eukaryotic chromosomes consist of large arrays of DNA repeats that undergo very rapid evolution. Nelumbo nucifera Gaertn. (sacred lotus) is a phylogenetic relict and an aquatic perennial basal eudicot. Studies concerning the centromeres of this basal eudicot species could provide ancient evolutionary perspectives. In this study, we characterized the centromeric marker protein NnCenH3 (sacred lotus centromere-specific histone H3 variant), and used a chromatin immunoprecipitation (ChIP)-based technique to recover the NnCenH3 nucleosome-associated sequences of sacred lotus. The properties of the centromere-binding protein and DNA sequences revealed notable divergence between sacred lotus and other flowering plants, including the following factors: (i) an NnCenH3 alternative splicing variant comprising only a partial centromere-targeting domain, (ii) active genes with low transcription levels in the NnCenH3 nucleosomal regions, and (iii) the prevalence of the Ty1/copia class of long terminal repeat (LTR) retrotransposons in the centromeres of sacred lotus chromosomes. In addition, the dynamic natures of the centromeric region showed that some of the centromeric repeat DNA sequences originated from telomeric repeats, and a pair of centromeres on the dicentric chromosome 1 was inactive in the metaphase cells of sacred lotus. Our characterization of the properties of centromeric DNA structure within the sacred lotus genome describes a centromeric profile in ancient basal eudicots and might provide evidence of the origins and evolution of centromeres. Furthermore, the identification of centromeric DNA sequences is of great significance for the assembly of the sacred lotus genome. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

Torsion of DNA modeled as a heterogeneous fluctuating rod

NASA Astrophysics Data System (ADS)

Argudo, David; Purohit, Prashant K.

2014-01-01

We discuss the statistical mechanics of a heterogeneous elastic rod with bending, twisting and stretching. Our model goes beyond earlier works where only homogeneous rods were considered in the limit of high forces and long lengths. Our methods allow us to consider shorter fluctuating rods for which boundary conditions can play an important role. We use our theory to study structural transitions in torsionally constrained DNA where there is coexistence of states with different effective properties. In particular, we examine whether a newly discovered left-handed DNA conformation called L-DNA is a mixture of two known states. We also use our model to investigate the mechanical effects of the binding of small molecules to DNA. For both these applications we make experimentally falsifiable predictions.

Atomically precise arrays of fluorescent silver clusters: a modular approach for metal cluster photonics on DNA nanostructures.

PubMed

Copp, Stacy M; Schultz, Danielle E; Swasey, Steven; Gwinn, Elisabeth G

2015-03-24

The remarkable precision that DNA scaffolds provide for arraying nanoscale optical elements enables optical phenomena that arise from interactions of metal nanoparticles, dye molecules, and quantum dots placed at nanoscale separations. However, control of ensemble optical properties has been limited by the difficulty of achieving uniform particle sizes and shapes. Ligand-stabilized metal clusters offer a route to atomically precise arrays that combine desirable attributes of both metals and molecules. Exploiting the unique advantages of the cluster regime requires techniques to realize controlled nanoscale placement of select cluster structures. Here we show that atomically monodisperse arrays of fluorescent, DNA-stabilized silver clusters can be realized on a prototypical scaffold, a DNA nanotube, with attachment sites separated by <10 nm. Cluster attachment is mediated by designed DNA linkers that enable isolation of specific clusters prior to assembly on nanotubes and preserve cluster structure and spectral purity after assembly. The modularity of this approach generalizes to silver clusters of diverse sizes and DNA scaffolds of many types. Thus, these silver cluster nano-optical elements, which themselves have colors selected by their particular DNA templating oligomer, bring unique dimensions of control and flexibility to the rapidly expanding field of nano-optics.

Hybridization assay of insect antifreezing protein gene by novel multilayered porous silicon nucleic acid biosensor.

PubMed

Lv, Xiaoyi; Chen, Liangliang; Zhang, Hongyan; Mo, Jiaqing; Zhong, Furu; Lv, Changwu; Ma, Ji; Jia, Zhenhong

2013-01-15

A fabrication of a novel simple porous silicon polybasic photonic crystal with symmetrical structure has been reported as a nucleic acid biosensor for detecting antifreeze protein gene in insects (Microdera puntipennis dzhungarica), which would be helpful in the development of some new transgenic plants with tolerance of freezing stress. Compared to various porous silicon-based photonic configurations, porous silicon polytype layered structure is quite easy to prepare and shows more stability; moreover, polybasic photonic crystals with symmetrical structure exhibit interesting optical properties with a sharp resonance in the reflectance spectrum, giving a higher Q factor which causes higher sensitivity for sensing performance. In this experiment, DNA oligonucleotides were immobilized into the porous silicon pores using a standard crosslink chemistry method. The porous silicon polybasic symmetrical structure sensor possesses high specificity in performing controlled experiments with non-complementary DNA. The detection limit was found to be 21.3nM for DNA oligonucleotides. The fabricated multilayered porous silicon-based DNA biosensor has potential commercial applications in clinical chemistry for determination of an antifreeze protein gene or other genes. Copyright © 2012 Elsevier B.V. All rights reserved.

Mechanical characteriztion of single-stranded DNA and single-walled carbon nanotube hybrid structures

NASA Astrophysics Data System (ADS)

Rokadia, Husein Juzer

Hybrid nanostructures of single-stranded DNA (ssDNA) and single-walled carbon nanotubes are being proposed as the basis for the next generation of biosensors. For such biosensors, mechanical properties such as the Young's modulus of the hybrid structures play a critical role, which to the best of the author's knowledge is still unknown. Thus, the determination of the Young's modulus of the ssDNA/swCNT hybrid structures was the primary objective of this study. Hybrid structures of 30mer polyT ssDNA and HiPCORTM swCNTs were conjugated using a well known non-covalent interaction protocol. Atomic force microscopy (AFM) was used to scan and generate topographic images and perform nanoindentation tests on the hybrid structures. Molecular dynamics (MD) simulations using a commercial MD program, Materials StudioRTM were performed to study the nature of non-covalent interactions between the ssDNA and the swCNT on the pico-second timescale. AFM topography scans of the bare control HiPCORTM swCNTs indicated an average diameter of about 1.0 nm and length of 800 nm. Similarly, the control 30mer polyT ssDNA was found to resemble a half-hemispherical domed structure with an average height of 2.1 nm. Nanoindentation tests yielded the transverse Young's modulus of the control swCNTs to be 78.0 GPa. The control ssDNA were found to have a Young's modulus of 3.3 GPa and 4.0 MPa in dry and wet environments, respectively. Topographic scans of the ssDNA/swCNT hybrid structures showed the slender swCNTs fully or partially coated along their lengths by ssDNA. The height of the hybrid structures ranged from 2.5 nm to 7.5 nm. Nanoindentation tests on the ssDNA coated portions of the hybrid structures indicated that, their Young's modulus exponentially decreased with increasing coating thickness. Thinly coated sections were found to have a Young's modulus of 100.0 GPa and 7.0 MPa in dry and wet conditions respectively. The thick walled hybrid sections were found to have an average Young's modulus of 4.5 GPa and 1.0 GPa in the dry and wet environments, respectively. MD results indicated that the wrapping of the ssDNA had a significant impact on the hybrid structures. The longitudinal Young's modulus of a hybrid structure was found to be approximately 50.0 GPa, compared to a bare nanotube whose Young's modulus was approximately 800 GPa. Overall, the experimental and numerical results displayed consistent trends. The experimental results reported the swCNTs to have the highest transverse Young's modulus followed by the hybrids and the ssDNA. Similarly, the numerical simulations predicted the highest longitudinal Young's modulus for the swCNTs, followed by the hybrids and the DNA.

Programmable disorder in random DNA tilings

NASA Astrophysics Data System (ADS)

Tikhomirov, Grigory; Petersen, Philip; Qian, Lulu

2017-03-01

Scaling up the complexity and diversity of synthetic molecular structures will require strategies that exploit the inherent stochasticity of molecular systems in a controlled fashion. Here we demonstrate a framework for programming random DNA tilings and show how to control the properties of global patterns through simple, local rules. We constructed three general forms of planar network—random loops, mazes and trees—on the surface of self-assembled DNA origami arrays on the micrometre scale with nanometre resolution. Using simple molecular building blocks and robust experimental conditions, we demonstrate control of a wide range of properties of the random networks, including the branching rules, the growth directions, the proximity between adjacent networks and the size distribution. Much as combinatorial approaches for generating random one-dimensional chains of polymers have been used to revolutionize chemical synthesis and the selection of functional nucleic acids, our strategy extends these principles to random two-dimensional networks of molecules and creates new opportunities for fabricating more complex molecular devices that are organized by DNA nanostructures.

Characterization of biochemical properties of Bacillus subtilis RecQ helicase.

PubMed

Qin, Wei; Liu, Na-Nv; Wang, Lijun; Zhou, Min; Ren, Hua; Bugnard, Elisabeth; Liu, Jie-Lin; Zhang, Lin-Hu; Vendôme, Jeremie; Hu, Jin-Shan; Xi, Xu Guang

2014-12-01

RecQ family helicases function as safeguards of the genome. Unlike Escherichia coli, the Gram-positive Bacillus subtilis bacterium possesses two RecQ-like homologues, RecQ[Bs] and RecS, which are required for the repair of DNA double-strand breaks. RecQ[Bs] also binds to the forked DNA to ensure a smooth progression of the cell cycle. Here we present the first biochemical analysis of recombinant RecQ[Bs]. RecQ[Bs] binds weakly to single-stranded DNA (ssDNA) and blunt-ended double-stranded DNA (dsDNA) but strongly to forked dsDNA. The protein exhibits a DNA-stimulated ATPase activity and ATP- and Mg(2+)-dependent DNA helicase activity with a 3' → 5' polarity. Molecular modeling shows that RecQ[Bs] shares high sequence and structure similarity with E. coli RecQ. Surprisingly, RecQ[Bs] resembles the truncated Saccharomyces cerevisiae Sgs1 and human RecQ helicases more than RecQ[Ec] with regard to its enzymatic activities. Specifically, RecQ[Bs] unwinds forked dsDNA and DNA duplexes with a 3'-overhang but is inactive on blunt-ended dsDNA and 5'-overhung duplexes. Interestingly, RecQ[Bs] unwinds blunt-ended DNA with structural features, including nicks, gaps, 5'-flaps, Kappa joints, synthetic replication forks, and Holliday junctions. We discuss these findings in the context of RecQ[Bs]'s possible functions in preserving genomic stability. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

DNA sequence determinants controlling affinity, stability and shape of DNA complexes bound by the nucleoid protein Fis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hancock, Stephen P.; Stella, Stefano; Cascio, Duilio

The abundant Fis nucleoid protein selectively binds poorly related DNA sequences with high affinities to regulate diverse DNA reactions. Fis binds DNA primarily through DNA backbone contacts and selects target sites by reading conformational properties of DNA sequences, most prominently intrinsic minor groove widths. High-affinity binding requires Fis-stabilized DNA conformational changes that vary depending on DNA sequence. In order to better understand the molecular basis for high affinity site recognition, we analyzed the effects of DNA sequence within and flanking the core Fis binding site on binding affinity and DNA structure. X-ray crystal structures of Fis-DNA complexes containing variable sequencesmore » in the noncontacted center of the binding site or variations within the major groove interfaces show that the DNA can adapt to the Fis dimer surface asymmetrically. We show that the presence and position of pyrimidine-purine base steps within the major groove interfaces affect both local DNA bending and minor groove compression to modulate affinities and lifetimes of Fis-DNA complexes. Sequences flanking the core binding site also modulate complex affinities, lifetimes, and the degree of local and global Fis-induced DNA bending. In particular, a G immediately upstream of the 15 bp core sequence inhibits binding and bending, and A-tracts within the flanking base pairs increase both complex lifetimes and global DNA curvatures. Taken together, our observations support a revised DNA motif specifying high-affinity Fis binding and highlight the range of conformations that Fis-bound DNA can adopt. Lastly, the affinities and DNA conformations of individual Fis-DNA complexes are likely to be tailored to their context-specific biological functions.« less

DNA sequence determinants controlling affinity, stability and shape of DNA complexes bound by the nucleoid protein Fis

DOE PAGES

Hancock, Stephen P.; Stella, Stefano; Cascio, Duilio; ...

2016-03-09

The abundant Fis nucleoid protein selectively binds poorly related DNA sequences with high affinities to regulate diverse DNA reactions. Fis binds DNA primarily through DNA backbone contacts and selects target sites by reading conformational properties of DNA sequences, most prominently intrinsic minor groove widths. High-affinity binding requires Fis-stabilized DNA conformational changes that vary depending on DNA sequence. In order to better understand the molecular basis for high affinity site recognition, we analyzed the effects of DNA sequence within and flanking the core Fis binding site on binding affinity and DNA structure. X-ray crystal structures of Fis-DNA complexes containing variable sequencesmore » in the noncontacted center of the binding site or variations within the major groove interfaces show that the DNA can adapt to the Fis dimer surface asymmetrically. We show that the presence and position of pyrimidine-purine base steps within the major groove interfaces affect both local DNA bending and minor groove compression to modulate affinities and lifetimes of Fis-DNA complexes. Sequences flanking the core binding site also modulate complex affinities, lifetimes, and the degree of local and global Fis-induced DNA bending. In particular, a G immediately upstream of the 15 bp core sequence inhibits binding and bending, and A-tracts within the flanking base pairs increase both complex lifetimes and global DNA curvatures. Taken together, our observations support a revised DNA motif specifying high-affinity Fis binding and highlight the range of conformations that Fis-bound DNA can adopt. Lastly, the affinities and DNA conformations of individual Fis-DNA complexes are likely to be tailored to their context-specific biological functions.« less

The architecture of neutrophil extracellular traps investigated by atomic force microscopy

NASA Astrophysics Data System (ADS)

Pires, Ricardo H.; Felix, Stephan B.; Delcea, Mihaela

2016-07-01

Neutrophils are immune cells that engage in a suicidal pathway leading to the release of partially decondensed chromatin, or neutrophil extracellular traps (NETs). NETs behave as a double edged sword; they can bind to pathogens thereby ensnaring them and limiting their spread during infection; however, they may bind to host circulating materials and trigger thrombotic events, and are associated with autoimmune disorders. Despite the fundamental role of NETs as part of an immune system response, there is currently a very poor understanding of how their nanoscale properties are reflected in their macroscopic impact. In this work, using a combination of fluorescence and atomic force microscopy, we show that NETs appear as a branching filament network that results in a substantially organized porous structure with openings with 0.03 +/- 0.04 μm2 on average and thus in the size range of small pathogens. Topological profiles typically up to 3 +/- 1 nm in height are compatible with a ``beads on a string'' model of nucleosome chromatin. Typical branch lengths of 153 +/- 103 nm appearing as rigid rods and height profiles of naked DNA in NETs of 1.2 +/- 0.5 nm are indicative of extensive DNA supercoiling throughout NETs. The presence of DNA duplexes could also be inferred from force spectroscopy and the occurrence of force plateaus that ranged from ~65 pN to 300 pN. Proteolytic digestion of NETs resulted in widespread disassembly of the network structure and considerable loss of mechanical properties. Our results suggest that the underlying structure of NETs is considerably organized and that part of its protein content plays an important role in maintaining its mesh architecture. We anticipate that NETs may work as microscopic mechanical sieves with elastic properties that stem from their DNA-protein composition, which is able to segregate particles also as a result of their size. Such a behavior may explain their participation in capturing pathogens and their association with thrombosis.Neutrophils are immune cells that engage in a suicidal pathway leading to the release of partially decondensed chromatin, or neutrophil extracellular traps (NETs). NETs behave as a double edged sword; they can bind to pathogens thereby ensnaring them and limiting their spread during infection; however, they may bind to host circulating materials and trigger thrombotic events, and are associated with autoimmune disorders. Despite the fundamental role of NETs as part of an immune system response, there is currently a very poor understanding of how their nanoscale properties are reflected in their macroscopic impact. In this work, using a combination of fluorescence and atomic force microscopy, we show that NETs appear as a branching filament network that results in a substantially organized porous structure with openings with 0.03 +/- 0.04 μm2 on average and thus in the size range of small pathogens. Topological profiles typically up to 3 +/- 1 nm in height are compatible with a ``beads on a string'' model of nucleosome chromatin. Typical branch lengths of 153 +/- 103 nm appearing as rigid rods and height profiles of naked DNA in NETs of 1.2 +/- 0.5 nm are indicative of extensive DNA supercoiling throughout NETs. The presence of DNA duplexes could also be inferred from force spectroscopy and the occurrence of force plateaus that ranged from ~65 pN to 300 pN. Proteolytic digestion of NETs resulted in widespread disassembly of the network structure and considerable loss of mechanical properties. Our results suggest that the underlying structure of NETs is considerably organized and that part of its protein content plays an important role in maintaining its mesh architecture. We anticipate that NETs may work as microscopic mechanical sieves with elastic properties that stem from their DNA-protein composition, which is able to segregate particles also as a result of their size. Such a behavior may explain their participation in capturing pathogens and their association with thrombosis. Electronic supplementary information (ESI) available: Fluorescence microscopy and AFM images of NETs; fluorescence and AFM images of chromatin of a neutrophil adhered on mica; height contrast AFM image of two plasmids evidencing supercoiling. See DOI: 10.1039/c6nr03416k

Long repeating (TTAGGG)n single stranded DNA self-condenses into compact beaded filaments stabilized by G-quadruplex formation.

PubMed

Kar, Anirban; Jones, Nathan; Arat, N Özlem; Fishel, Richard; Griffith, Jack

2018-04-19

Conformations adopted by long stretches of single stranded DNA (ssDNA) are of central interest in understanding the architecture of replication forks, R loops, and other structures generated during DNA metabolism in vivo. This is particularly so if the ssDNA consists of short nucleotide repeats. Such studies have been hampered by the lack of defined substrates greater than ~150 nt, and the absence of high-resolution biophysical approaches. Here we describe the generation of very long ssDNA consisting of the mammalian telomeric repeat (5'-TTAGGG-3')n as well as the interrogation of its structure by electron microscopy (EM) and single molecule magnetic tweezers (smMT). This repeat is of particular interest as it contains a run of 3 contiguous guanine residues capable of forming G quartets as ssDNA. Fluorescent-dye exclusion assays confirmed that this G-strand ssDNA forms ubiquitous G-quadruplex folds. EM revealed thick bead-like filaments that condensed the DNA ~12 fold. The bead-like structures were 5 nm and 8 nm in diameter and linked by thin filaments. The G-strand ssDNA displayed initial stability to smMT force extension that ultimately released in steps that were multiples ~28 nm at forces between 6-12 pN; well below the >20 pN required to unravel G-quadruplexes. Most smMT steps were consistent with the disruption of the beads seen by EM. Binding by RAD51 distinctively altered the force extension properties of the G-strand ssDNA, suggesting a stochastic G-quadruplex-dependent condensation model that is discussed. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

Improved DNA hybridization parameters by Twisted Intercalating Nucleic Acid (TINA).

PubMed

Schneider, Uffe Vest

2012-01-01

This thesis establishes oligonucleotide design rules and applications of a novel group of DNA stabilizing molecules collectively called Twisted Intercalating Nucleic Acid - TINA. Three peer-reviewed publications form the basis for the thesis. One publication describes an improved and rapid method for determination of DNA melting points and two publications describe the effects of positioning TINA molecules in parallel triplex helix and antiparallel duplex helix forming DNA structures. The third publication establishes that TINA molecules containing oligonucleotides improve an antiparallel duplex hybridization based capture assay's analytical sensitivity compared to conventionel DNA oligonucleotides. Clinical microbiology is traditionally based on pathogenic microorganisms' culture and serological tests. The introduction of DNA target amplification methods like PCR has improved the analytical sensitivity and total turn around time involved in clinical diagnostics of infections. Due to the relatively weak hybridization between the two strands of double stranded DNA, a number of nucleic acid stabilizing molecules have been developed to improve the sensitivity of DNA based diagnostics through superior binding properties. A short introduction is given to Watson-Crick and Hoogsteen based DNA binding and the derived DNA structures. A number of other nucleic acid stabilizing molecules are described. The stabilizing effect of TINA molecules on different DNA structures is discussed and considered in relation to other nucleic acid stabilizing molecules and in relation to future use of TINA containing oligonucleotides in clinical diagnostics and therapy. In conclusion, design of TINA modified oligonucleotides for antiparallel duplex helixes and parallel triplex helixes follows simple purpose dependent rules. TINA molecules are well suited for improving multiplex PCR assays and can be used as part of novel technologies. Future research should test whether combinations of TINA molecules and other nucleic acid stabilizing molecules can increase analytical sensitivity whilst maintaining nucleobase mismatch discrimination in triplex helix based diagnostic assays.

Sequence-dependent modelling of local DNA bending phenomena: curvature prediction and vibrational analysis.

PubMed

Vlahovicek, K; Munteanu, M G; Pongor, S

1999-01-01

Bending is a local conformational micropolymorphism of DNA in which the original B-DNA structure is only distorted but not extensively modified. Bending can be predicted by simple static geometry models as well as by a recently developed elastic model that incorporate sequence dependent anisotropic bendability (SDAB). The SDAB model qualitatively explains phenomena including affinity of protein binding, kinking, as well as sequence-dependent vibrational properties of DNA. The vibrational properties of DNA segments can be studied by finite element analysis of a model subjected to an initial bending moment. The frequency spectrum is obtained by applying Fourier analysis to the displacement values in the time domain. This analysis shows that the spectrum of the bending vibrations quite sensitively depends on the sequence, for example the spectrum of a curved sequence is characteristically different from the spectrum of straight sequence motifs of identical basepair composition. Curvature distributions are genome-specific, and pronounced differences are found between protein-coding and regulatory regions, respectively, that is, sites of extreme curvature and/or bendability are less frequent in protein-coding regions. A WWW server is set up for the prediction of curvature and generation of 3D models from DNA sequences (http:@www.icgeb.trieste.it/dna).

Investigation of biochemical property changes in activation-induced CD 8 + T cell apoptosis using Raman spectroscopy

NASA Astrophysics Data System (ADS)

Lee, Young Ju; Ahn, Hyung Joon; Lee, Gi-Ja; Jung, Gyeong Bok; Lee, Gihyun; Kim, Dohyun; Shin, Jae-Ho; Jin, Kyung-Hyun; Park, Hun-Kuk

2015-07-01

The study was to investigate the changes in biochemical properties of activated mature CD8+ T cells related to apoptosis at a molecular level. We confirmed the activation and apoptosis of CD8+ T cells by fluorescence-activated cell sorting and atomic force microscopy and then performed Raman spectral measurements on activated mature CD8+ T cells and cellular deoxyribose nucleic acid (DNA). In the activated mature CD8+ T cells, there were increases in protein spectra at 1002 and 1234 cm-1. In particular, to assess the apoptosis-related DNA spectral signatures, we investigated the spectra of the cellular DNA isolated from resting and activated mature CD8+ T cells. Raman spectra at 765 to 786 cm-1 and 1053 to 1087 cm-1 were decreased in activated mature DNA. In addition, we analyzed Raman spectrum using the multivariate statistical method including principal component analysis. Raman spectra of activated mature DNA are especially well-discriminated from those of resting DNA. Our findings regarding the biochemical and structural changes associated with apoptosis in activated mature T cells and cellular DNA according to Raman spectroscopy provide important insights into allospecific immune responses generated after organ transplantation, and may be useful for therapeutic manipulation of the immune response.

DNA Polymerase Eta and Chemotherapeutic Agents

PubMed Central

2011-01-01

Abstract The discovery of human DNA polymerase eta (pol η) has a major impact on the fields of DNA replication/repair fields. Since the discovery of human pol η, a number of new DNA polymerases with the ability to bypass various DNA lesions have been discovered. Among these polymerases, pol η is the most extensively studied lesion bypass polymerase with a defined major biological function, that is, to replicate across the cyclobutane pyrimidine dimers introduced by UV irradiation. Cyclobutane pyrimidine dimer is a major DNA lesion that causes distortion of DNA structure and block the replicative DNA polymerases during DNA replication process. Genetic defects in the pol η gene, Rad30, results in a disease called xeroderma pigmentosum variant. This review focuses on the overall properties of pol η and the mechanism that involved in regulating its activity in cells. In addition, the role of pol η in the action of DNA-targeting anticancer compounds is also discussed. Antioxid. Redox Signal. 14, 2521–2529. PMID:21050139

Synthesis, characterization, crystal structure and theoretical study of a compound with benzodiazole ring: antimicrobial activity and DNA binding.

PubMed

Latha, P; Kodisundaram, P; Sundararajan, M L; Jeyakumar, T

2014-08-14

2-(Thiophen-2-yl)-1-((thiophen-2-yl)methyl)-1H-1,3-benzodiazole (HL) is synthesized and characterized by elemental analysis, UV-Vis, FT-IR, (1)H, (13)C NMR, mass spectra, scanning electron microscope (SEM) and single crystal X-ray diffraction. The crystal structure is stabilized by intermolecular CH⋯N and CH⋯π interactions. The molecular structure is also optimized at the B3LYP/6-31G level using density functional theory (DFT). The structural parameters from the theory are nearer to those of crystal, the calculated total energy of coordination is -1522.814a.u. The energy of HOMO-LUMO and the energy gap are -0.20718, -0.04314, 0.16404a.u, respectively. All data obtained from the spectral studies support the structural properties of the compound HL. The benzimidazole ring is essentially planar. The in vitro biological screening effects of the synthesized compound is tested against four bacterial and four fungal strains by well diffusion method. Antioxidant property and DNA binding behaviour of the compound has been investigated using spectrophotometric method. Copyright © 2014 Elsevier B.V. All rights reserved.

Self-assembling DNA nanotubes to connect molecular landmarks

NASA Astrophysics Data System (ADS)

Mohammed, Abdul M.; Šulc, Petr; Zenk, John; Schulman, Rebecca

2017-05-01

Within cells, nanostructures are often organized using local assembly rules that produce long-range order. Because these rules can take into account the cell's current structure and state, they can enable complexes, organelles or cytoskeletal structures to assemble around existing cellular components to form architectures. Although many methods for self-assembling biomolecular nanostructures have been developed, few can be programmed to assemble structures whose form depends on the identity and organization of structures already present in the environment. Here, we demonstrate that DNA nanotubes can grow to connect pairs of molecular landmarks with different separation distances and relative orientations. DNA tile nanotubes nucleate at these landmarks and grow while their free ends diffuse. The nanotubes can then join end to end to form stable connections, with unconnected nanotubes selectively melted away. Connections form between landmark pairs separated by 1-10 µm in more than 75% of cases and can span a surface or three dimensions. This point-to-point assembly process illustrates how self-assembly kinetics can be designed to produce structures with a desired physical property rather than a specific shape.

Macroscopic modeling and simulations of supercoiled DNA with bound proteins

NASA Astrophysics Data System (ADS)

Huang, Jing; Schlick, Tamar

2002-11-01

General methods are presented for modeling and simulating DNA molecules with bound proteins on the macromolecular level. These new approaches are motivated by the need for accurate and affordable methods to simulate slow processes (on the millisecond time scale) in DNA/protein systems, such as the large-scale motions involved in the Hin-mediated inversion process. Our approaches, based on the wormlike chain model of long DNA molecules, introduce inhomogeneous potentials for DNA/protein complexes based on available atomic-level structures. Electrostatically, treat those DNA/protein complexes as sets of effective charges, optimized by our discrete surface charge optimization package, in which the charges are distributed on an excluded-volume surface that represents the macromolecular complex. We also introduce directional bending potentials as well as non-identical bead hydrodynamics algorithm to further mimic the inhomogeneous effects caused by protein binding. These models thus account for basic elements of protein binding effects on DNA local structure but remain computational tractable. To validate these models and methods, we reproduce various properties measured by both Monte Carlo methods and experiments. We then apply the developed models to study the Hin-mediated inversion system in long DNA. By simulating supercoiled, circular DNA with or without bound proteins, we observe significant effects of protein binding on global conformations and long-time dynamics of the DNA on the kilo basepair length.

Effects of polyamines and methylglyoxal bis(guanylhydrazone) on hepatic nuclear structure and deoxyribonucleic acid template activity.

PubMed Central

Brown, K B; Nelson, N F; Brown, D G

1975-01-01

1. The interaction of polyamines and methylglyoxal bis(guanythydrazone) (1, 1'-[(methylethanediylidene)-dinitrilo]diguanidine) with isolated rat liver nuclei was investigated by electron microscopy. 2. At 4mM, putrescine was without effect; however, spermidine, spermine or methylglyoxal bis(guanythydrazone) resulted in dispersed chromatin and alterations in nucleolar structure. In addition, spermidine or methylglyoxal bis(guanylhydrazone) caused marked aggregation of interchromatin granules. 3. The DNA template property of calf thymus DNA was examined by using DNA polymerases from Escherichia coli, Micrococcus lysodeikticus and calf thymus in the presence of 0-5 mM-amine. 4. In the presence of DNA polymerase, spermine or methylglyoxal bis(guanylhydrazone) inhibited activity, whereas putrescine or spermidine had much less effect or in some cases stimulated [3H]dTMP incorporation. 5. Template activity which was inhibited by spermine or methylglyoxal bis(guanylhydrazone) could be partially restored by additional DNA or enzyme. 6. When mixed with calf thymus DNA, calf thymus histone inhibited template activity as measured with E. coli DNA polymerase. The template activity of such a 'histone-nucleate' could not be restored by putrescine, spermidine, spermine or methylglyoxal bis(guanylhydrazone). 7. DNA template activity of isolated rat liver nuclei was tested by using E. coli DNA polymerase. None of the amines was able to increase the template activity of the nuclear DNA in vitro. Images PLATE 1 PMID:1218090

Effects of polyamines and methylglyoxal bis(guanylhydrazone) on hepatic nuclear structure and deoxyribonucleic acid template activity.

PubMed

Brown, K B; Nelson, N F; Brown, D G

1975-12-01

1. The interaction of polyamines and methylglyoxal bis(guanythydrazone) (1, 1'-[(methylethanediylidene)-dinitrilo]diguanidine) with isolated rat liver nuclei was investigated by electron microscopy. 2. At 4mM, putrescine was without effect; however, spermidine, spermine or methylglyoxal bis(guanythydrazone) resulted in dispersed chromatin and alterations in nucleolar structure. In addition, spermidine or methylglyoxal bis(guanylhydrazone) caused marked aggregation of interchromatin granules. 3. The DNA template property of calf thymus DNA was examined by using DNA polymerases from Escherichia coli, Micrococcus lysodeikticus and calf thymus in the presence of 0-5 mM-amine. 4. In the presence of DNA polymerase, spermine or methylglyoxal bis(guanylhydrazone) inhibited activity, whereas putrescine or spermidine had much less effect or in some cases stimulated [3H]dTMP incorporation. 5. Template activity which was inhibited by spermine or methylglyoxal bis(guanylhydrazone) could be partially restored by additional DNA or enzyme. 6. When mixed with calf thymus DNA, calf thymus histone inhibited template activity as measured with E. coli DNA polymerase. The template activity of such a 'histone-nucleate' could not be restored by putrescine, spermidine, spermine or methylglyoxal bis(guanylhydrazone). 7. DNA template activity of isolated rat liver nuclei was tested by using E. coli DNA polymerase. None of the amines was able to increase the template activity of the nuclear DNA in vitro.

DNA bases ring-expanded with a cyclopentadiene free radical: a theoretical investigation of building blocks with diradical character.

PubMed

Zhao, Peiwen; Bu, Yuxiang

2016-01-14

In this work, we computationally design radical nucleobases which possess improved electronic properties, especially diradical properties through introducing a cyclopentadiene radical. We predict that the detailed electromagnetic features of base assemblies are based on the orientation of the extra five-membered cyclopentadiene ring. Broken symmetry DFT calculations take into account the relevant structures and properties. Our results reveal that both the radicalized DNA bases and the base pairs formed when they combine with their counterparts remain stable and display larger spin delocalization. The mode of embedding the cyclopentadiene free radical in the structures has some influence on the degree of π-conjugation, which results in various diradical characteristics. Single-layered radical base pairs all have an open-shell singlet ground state, but the energy difference between singlet and triplet is not significant. For two-layered radical base pairs, the situation is more complex. All of them have an open-shell state as their ground state, including an open-shell singlet state and an open-shell triplet state. That is, the majority of radical base pairs possess anti-ferromagnetic or ferromagnetic characteristics. We present here a more in-depth discussion and analyses to study the magnetic characteristics of radical bases and base pairs. As an important factor, two-layered radical base pairs also have been carefully analyzed. We hope that all the measurements and results presented here will stimulate further detailed insights into the related mechanisms in modified DNA bases and the design of better ring-expanded DNA magnetic materials.

Subunit Conformations and Assembly States of a DNA Translocating Motor: The Terminase of Bacteriophage P22

PubMed Central

Němeček, Daniel; Gilcrease, Eddie B.; Kang, Sebyung; Prevelige, Peter E.; Casjens, Sherwood; Thomas, George J.

2007-01-01

Bacteriophage P22, a podovirus infecting strains of Salmonella typhimurium, packages a 42 kbp genome using a headful mechanism. DNA translocation is accomplished by the phage terminase, a powerful molecular motor consisting of large and small subunits. Although many of the structural proteins of the P22 virion have been well characterized, little is known about the terminase subunits and their molecular mechanism of DNA translocation. We report here structural and assembly properties of ectopically expressed and highly purified terminase large and small subunits. The large subunit (gp2), which contains the nuclease and ATPase activities of terminase, exists as a stable monomer with an α/β fold. The small subunit (gp3), which recognizes DNA for packaging and may regulate gp2 activity, exhibits a highly α-helical secondary structure and self-associates to form a stable oligomeric ring in solution. For wildtype gp3, the ring contains nine subunits, as demonstrated by hydrodynamic measurements, electron microscopy and native mass spectrometry. We have also characterized a gp3 mutant (Ala 112 → Thr) that forms a ten subunit ring, despite a subunit fold indistinguishable from wildtype. Both the nonameric and decameric gp3 rings exhibit nonspecific DNA binding activity, and gp2 is able to bind strongly to the DNA/gp3 complex but not to DNA alone. We propose a scheme for the roles of P22 terminase large and small subunits in the recruitment and packaging of viral DNA and discuss the model in relation to proposals for terminase-driven DNA translocation in other phages. PMID:17945256

Anisotropic nanomaterials: Synthesis, optical and magnetic properties, and applications

NASA Astrophysics Data System (ADS)

Banholzer, Matthew John

As nanoscience and nanotechnology mature, anisotropic metal nanostructures are emerging in a variety of contexts as valuable class of nanostructures due to their distinctive attributes. With unique properties ranging from optical to magnetic and beyond, these structures are useful in many new applications. Chapter two discusses the nanodisk code: a linear array of metal disk pairs that serve as surface-enhanced Raman scattering substrates. These multiplexing structures employ a binary encoding scheme, perform better than previous nanowires designs (in the context of SERS) and are useful for both convert encoding and tagging of substrates (based both on spatial disk position and spectroscopic response) as well as biomolecule detection (e.g. DNA). Chapter three describes the development of improved, silver-based nanodisk code structures. Work was undertaken to generate structures with high yield and reproducibility and to reoptimize the geometry of each disk pair for maximum Raman enhancement. The improved silver structures exhibit greater enhancement than Au structures (leading to lower DNA detection limits), convey additional flexibility, and enable trinary encoding schemes where far more unique structures can be created. Chapter four considers the effect of roughness on the plasmonic properties of nanorod structures and introduces a novel method to smooth the end-surfaces of nanorods structures. The smoothing technique is based upon a two-step process relying upon diffusion control during nanowires growth and selective oxidation after each step of synthesis is complete. Empirical and theoretical work show that smoothed nanostructures have superior and controllable optical properties. Chapter five concerns silica-encapsulated gold nanoprisms. This encapsulation allows these highly sensitive prisms to remain stable and protected in solution, enabling their use as class-leading sensors. Theoretical study complements the empirical work, exploring the effect of encapsulation on the SPR of these structures. Chapter six focuses on the magnetic properties of Au-Ni heterostructures. In addition to demonstration of nanoconfinement effects based upon the anisotropy of the nanorods/nanodisk structure, the magnetic coupling of rod-disk heterostructures is examined. Subsequent investigations suggest that the magnetic behavior of disks can be influenced by nearby rod segments, leading to the creation of a three-state spin system that may prove useful in device applications.

Fabrication, characterization, and biological assessment of multilayer laminin γ2 DNA coatings on titanium surfaces.

PubMed

Yang, Guoli; Zhang, Jing; Dong, Wenjing; Liu, Li; Shi, Jue; Wang, Huiming

2016-03-21

The purpose of this work was to fabricate a multilayer laminin γ2 DNA coating on a titanium surface and evaluate its biological properties. A multilayer laminin γ2 DNA coating was fabricated on titanium using a layer-by-layer assembly technique. The rate of coating degradation was evaluated by detecting the amount of cDNA remaining. Surface analysis using X-ray photoelectron spectroscopy, atomic force microscopy, and surface contact angle measurements revealed the multilayer structure to consist of cationic lipid and confirmed that a laminin γ2 DNA layer could be fabricated on titanium via the layer-by-layer assembly process. The transfection efficiency was highest for five layers in the multilayer structure. HEK293 cells cultured on the multilayer films displayed significantly higher adhesion activity than the control group. The expression of laminin γ2 and the co-localization of integrin β4 and plectin were more obvious in HN4 cells cultured on the multilayer laminin γ2 DNA coating, while weak immunoreactivities were observed in the control group. We concluded that the DNA-loaded multilayer provided a surface with good biocompatibility and that the multilayer laminin γ2 DNA coating might be effective in improving cell adhesion and the formation of hemidesmosomes on titanium surfaces.

Fabrication, characterization, and biological assessment of multilayer laminin γ2 DNA coatings on titanium surfaces

PubMed Central

Yang, Guoli; Zhang, Jing; Dong, Wenjing; Liu, Li; Shi, Jue; Wang, Huiming

2016-01-01

The purpose of this work was to fabricate a multilayer laminin γ2 DNA coating on a titanium surface and evaluate its biological properties. A multilayer laminin γ2 DNA coating was fabricated on titanium using a layer-by-layer assembly technique. The rate of coating degradation was evaluated by detecting the amount of cDNA remaining. Surface analysis using X-ray photoelectron spectroscopy, atomic force microscopy, and surface contact angle measurements revealed the multilayer structure to consist of cationic lipid and confirmed that a laminin γ2 DNA layer could be fabricated on titanium via the layer-by-layer assembly process. The transfection efficiency was highest for five layers in the multilayer structure. HEK293 cells cultured on the multilayer films displayed significantly higher adhesion activity than the control group. The expression of laminin γ2 and the co-localization of integrin β4 and plectin were more obvious in HN4 cells cultured on the multilayer laminin γ2 DNA coating, while weak immunoreactivities were observed in the control group. We concluded that the DNA-loaded multilayer provided a surface with good biocompatibility and that the multilayer laminin γ2 DNA coating might be effective in improving cell adhesion and the formation of hemidesmosomes on titanium surfaces. PMID:26996815

Fabrication, characterization, and biological assessment of multilayer laminin γ2 DNA coatings on titanium surfaces

NASA Astrophysics Data System (ADS)

Yang, Guoli; Zhang, Jing; Dong, Wenjing; Liu, Li; Shi, Jue; Wang, Huiming

2016-03-01

The purpose of this work was to fabricate a multilayer laminin γ2 DNA coating on a titanium surface and evaluate its biological properties. A multilayer laminin γ2 DNA coating was fabricated on titanium using a layer-by-layer assembly technique. The rate of coating degradation was evaluated by detecting the amount of cDNA remaining. Surface analysis using X-ray photoelectron spectroscopy, atomic force microscopy, and surface contact angle measurements revealed the multilayer structure to consist of cationic lipid and confirmed that a laminin γ2 DNA layer could be fabricated on titanium via the layer-by-layer assembly process. The transfection efficiency was highest for five layers in the multilayer structure. HEK293 cells cultured on the multilayer films displayed significantly higher adhesion activity than the control group. The expression of laminin γ2 and the co-localization of integrin β4 and plectin were more obvious in HN4 cells cultured on the multilayer laminin γ2 DNA coating, while weak immunoreactivities were observed in the control group. We concluded that the DNA-loaded multilayer provided a surface with good biocompatibility and that the multilayer laminin γ2 DNA coating might be effective in improving cell adhesion and the formation of hemidesmosomes on titanium surfaces.

BIOPHYSICAL PROPERTIES OF NUCLEIC ACIDS AT SURFACES RELEVANT TO MICROARRAY PERFORMANCE.

PubMed

Rao, Archana N; Grainger, David W

2014-04-01

Both clinical and analytical metrics produced by microarray-based assay technology have recognized problems in reproducibility, reliability and analytical sensitivity. These issues are often attributed to poor understanding and control of nucleic acid behaviors and properties at solid-liquid interfaces. Nucleic acid hybridization, central to DNA and RNA microarray formats, depends on the properties and behaviors of single strand (ss) nucleic acids (e.g., probe oligomeric DNA) bound to surfaces. ssDNA's persistence length, radius of gyration, electrostatics, conformations on different surfaces and under various assay conditions, its chain flexibility and curvature, charging effects in ionic solutions, and fluorescent labeling all influence its physical chemistry and hybridization under assay conditions. Nucleic acid (e.g., both RNA and DNA) target interactions with immobilized ssDNA strands are highly impacted by these biophysical states. Furthermore, the kinetics, thermodynamics, and enthalpic and entropic contributions to DNA hybridization reflect global probe/target structures and interaction dynamics. Here we review several biophysical issues relevant to oligomeric nucleic acid molecular behaviors at surfaces and their influences on duplex formation that influence microarray assay performance. Correlation of biophysical aspects of single and double-stranded nucleic acids with their complexes in bulk solution is common. Such analysis at surfaces is not commonly reported, despite its importance to microarray assays. We seek to provide further insight into nucleic acid-surface challenges facing microarray diagnostic formats that have hindered their clinical adoption and compromise their research quality and value as genomics tools.

Theory of electron transfer and molecular state in DNA

NASA Astrophysics Data System (ADS)

Endres, Robert Gunter

2002-09-01

In this thesis, a mechanism for long-range electron transfer in DNA and a systematic search for high conductance DNA are developed. DNA is well known for containing the genetic code of all living species. On the other hand, there are some experimental indications that DNA can mediate effectively long-range electron transfer leading to the concept of chemistry at a distance. This can be important for DNA damage and healing. In the first part of the thesis, a possible mechanism for long-range electron transfer is introduced. The weak distance dependent electron transfer was experimentally observed using transition metal intercalators for donor and acceptor. In our model calculations, the transfer is mediated by the molecular analogue of a Kondo bound state well known from solid state physics of mixed-valence rare-earth compounds. We believe this is quite realistic, since localized d orbitals of the transition metal ions could function as an Anderson impurity embedded in a reservoir of rather delocalized molecular orbitals of the intercalator ligands and DNA pi orbitals. The effective Anderson model is solved with a physically intuitive variational ansatz as well as with the essentially exact DMRG method. The electronic transition matrix element, which is important because it contains the donor-acceptor distance dependence, is obtained with the Mulliken-Hush algorithm as well as from Born-Oppenheimer potential energy surfaces. Our possible explanation of long-range electron transfer is put in context to other more conventional mechanisms which also could lead to similar behavior. Another important issue of DNA is its possible use for nano-technology. Although DNA's mechanical properties are excellent, the question whether it can be conducting and be used for nano-wires is highly controversial. Experimentally, DNA shows conducting, semi-conducting and insulating properties. Motivated by these wide ranging experimental results on the conductivity of DNA, we have embarked on a theoretical effort to ascertain what conditions might induce such remarkable behavior. We use a combination of an ab initio density functional theory method and a parameterized Huckel-Slater-Koster model. Our focus here is to examine whether any likely DNA structures or environments can yield reduced activation gaps to conduction or enhanced electronic overlaps. In particular, we study a hypothetical stretched ribbon structure, A-, and B-form DNA, and the effects of counterions and humidity. Unlike solids, DNA and other molecules are considered soft condensed matter. Hence, we study the influence of vibrations upon the electronic structure of DNA. We calculate parameters for charge transfer rates between adjacent bases. We find good agreement between our estimated rates and recent experimental data assuming that torsional vibrations limit the charge transfer most significantly.

Computational strategies to address chromatin structure problems

NASA Astrophysics Data System (ADS)

Perišić, Ognjen; Schlick, Tamar

2016-06-01

While the genetic information is contained in double helical DNA, gene expression is a complex multilevel process that involves various functional units, from nucleosomes to fully formed chromatin fibers accompanied by a host of various chromatin binding enzymes. The chromatin fiber is a polymer composed of histone protein complexes upon which DNA wraps, like yarn upon many spools. The nature of chromatin structure has been an open question since the beginning of modern molecular biology. Many experiments have shown that the chromatin fiber is a highly dynamic entity with pronounced structural diversity that includes properties of idealized zig-zag and solenoid models, as well as other motifs. This diversity can produce a high packing ratio and thus inhibit access to a majority of the wound DNA. Despite much research, chromatin’s dynamic structure has not yet been fully described. Long stretches of chromatin fibers exhibit puzzling dynamic behavior that requires interpretation in the light of gene expression patterns in various tissue and organisms. The properties of chromatin fiber can be investigated with experimental techniques, like in vitro biochemistry, in vivo imagining, and high-throughput chromosome capture technology. Those techniques provide useful insights into the fiber’s structure and dynamics, but they are limited in resolution and scope, especially regarding compact fibers and chromosomes in the cellular milieu. Complementary but specialized modeling techniques are needed to handle large floppy polymers such as the chromatin fiber. In this review, we discuss current approaches in the chromatin structure field with an emphasis on modeling, such as molecular dynamics and coarse-grained computational approaches. Combinations of these computational techniques complement experiments and address many relevant biological problems, as we will illustrate with special focus on epigenetic modulation of chromatin structure.

DNA origami applications in cancer therapy.

PubMed

Udomprasert, Anuttara; Kangsamaksin, Thaned

2017-08-01

Due to the complexity and heterogeneity of cancer, the development of cancer diagnosis and therapy is still progressing, and a complete understanding of cancer biology remains elusive. Recently, cancer nanomedicine has gained much interest as a promising diagnostic and therapeutic strategy, as a wide range of nanomaterials possess unique physical properties that can render drug delivery systems safer and more effective. Also, targeted drug delivery and precision medicine have now become a new paradigm in cancer therapy. With nanocarriers, chemotherapeutic drugs could be directly delivered into target cancer cells, resulting in enhanced efficiency with fewer side-effects. DNA, a biomolecule with molecular self-assembly properties, has emerged as a versatile nanomaterial to construct multifunctional platforms; DNA nanostructures can be modified with functional groups to improve their utilities as biosensors or drug carriers. Such applications have become possible with the advent of the scaffolded DNA origami method. This breakthrough technique in structural DNA nanotechnology provides an easier and faster way to construct DNA nanostructures with various shapes. Several experiments proved that DNA origami nanostructures possess abilities to enhance efficacies of chemotherapy, reduce adverse side-effects, and even circumvent drug resistance. Here, we highlight the principles of the DNA origami technique and its applications in cancer therapeutics and discuss current challenges and opportunities to improve cancer detection and targeted drug delivery. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

The fluorescently responsive 3-(naphthalen-1-ylethynyl)-3-deaza-2'-deoxyguanosine discriminates cytidine via the DNA minor groove.

PubMed

Suzuki, Azusa; Yanagi, Masaki; Takeda, Takuya; Hudson, Robert H E; Saito, Yoshio

2017-09-26

A new environmentally responsive fluorescent nucleoside, 3-(naphthalen-1-ylethynyl)-3-deaza-2'-deoxyguanosine ( 3nz G), has been synthesized. The nucleoside, 3nz G, exhibited solvatochromic properties and when introduced into ODN probes it was able to recognize 2'-deoxycytidine in target strands by a distinct change in its emission wavelength through probing microenvironmental changes in the DNA minor groove. Thus, 3nz G has the potential for use as a fluorescent probe molecule for micro-structural studies of nucleic acids including the detection of single-base alterations in target DNA sequences.

Modeling of DNA local parameters predicts encrypted architectural motifs in Xenopus laevis ribosomal gene promoter.

PubMed

Roux-Rouquie, M; Marilley, M

2000-09-15

We have modeled local DNA sequence parameters to search for DNA architectural motifs involved in transcription regulation and promotion within the Xenopus laevis ribosomal gene promoter and the intergenic spacer (IGS) sequences. The IGS was found to be shaped into distinct topological domains. First, intrinsic bends split the IGS into domains of common but different helical features. Local parameters at inter-domain junctions exhibit a high variability with respect to intrinsic curvature, bendability and thermal stability. Secondly, the repeated sequence blocks of the IGS exhibit right-handed supercoiled structures which could be related to their enhancer properties. Thirdly, the gene promoter presents both inherent curvature and minor groove narrowing which may be viewed as motifs of a structural code for protein recognition and binding. Such pre-existing deformations could simply be remodeled during the binding of the transcription complex. Alternatively, these deformations could pre-shape the promoter in such a way that further remodeling is facilitated. Mutations shown to abolish promoter curvature as well as intrinsic minor groove narrowing, in a variant which maintained full transcriptional activity, bring circumstantial evidence for structurally-preorganized motifs in relation to transcription regulation and promotion. Using well documented X. laevis rDNA regulatory sequences we showed that computer modeling may be of invaluable assistance in assessing encrypted architectural motifs. The evidence of these DNA topological motifs with respect to the concept of structural code is discussed.

Conformational Dynamics of Mechanically Compliant DNA Nanostructures from Coarse-Grained Molecular Dynamics Simulations.

PubMed

Shi, Ze; Castro, Carlos E; Arya, Gaurav

2017-05-23

Structural DNA nanotechnology, the assembly of rigid 3D structures of complex yet precise geometries, has recently been used to design dynamic, mechanically compliant nanostructures with tunable equilibrium conformations and conformational distributions. Here we use coarse-grained molecular dynamics simulations to provide insights into the conformational dynamics of a set of mechanically compliant DNA nanostructures-DNA hinges that use single-stranded DNA "springs" to tune the equilibrium conformation of a layered double-stranded DNA "joint" connecting two stiff "arms" constructed from DNA helix bundles. The simulations reproduce the experimentally measured equilibrium angles between hinge arms for a range of hinge designs. The hinges are found to be structurally stable, except for some fraying of the open ends of the DNA helices comprising the hinge arms and some loss of base-pairing interactions in the joint regions coinciding with the crossover junctions, especially in hinges designed to exhibit a small bending angle that exhibit large local stresses resulting in strong kinks in their joints. Principal component analysis reveals that while the hinge dynamics are dominated by bending motion, some twisting and sliding of hinge arms relative to each other also exists. Forced deformation of the hinges reveals distinct bending mechanisms for hinges with short, inextensible springs versus those with longer, more extensible springs. Lastly, we introduce an approach for rapidly predicting equilibrium hinge angles from individual force-deformation behaviors of its single- and double-stranded DNA components. Taken together, these results demonstrate that coarse-grained modeling is a promising approach for designing, predicting, and studying the dynamics of compliant DNA nanostructures, where conformational fluctuations become important, multiple deformation mechanisms exist, and continuum approaches may not yield accurate properties.

Binding mechanism of PicoGreen to DNA characterized by magnetic tweezers and fluorescence spectroscopy.

PubMed

Wang, Ying; Schellenberg, Helene; Walhorn, Volker; Toensing, Katja; Anselmetti, Dario

2017-09-01

Fluorescent dyes are broadly used in many biotechnological applications to detect and visualize DNA molecules. However, their binding to DNA alters the structural and nanomechanical properties of DNA and, thus, interferes with associated biological processes. In this work we employed magnetic tweezers and fluorescence spectroscopy to investigate the binding of PicoGreen to DNA at room temperature in a concentration-dependent manner. PicoGreen is an ultrasensitive quinolinium nucleic acid stain exhibiting hardly any background signal from unbound dye molecules. By means of stretching and overwinding single, torsionally constrained, nick-free double-stranded DNA molecules, we acquired force-extension and supercoiling curves which allow quantifying DNA contour length, persistence length and other thermodynamical binding parameters, respectively. The results of our magnetic tweezers single-molecule binding study were well supported through analyzing the fluorescent spectra of stained DNA. On the basis of our work, we could identify a concentration-dependent bimodal binding behavior, where, apparently, PicoGreen associates to DNA as an intercalator and minor-groove binder simultaneously.

An Insilico Design of Nanoclay Based Nanocomposites and Scaffolds in Bone Tissue Engineering

NASA Astrophysics Data System (ADS)

Sharma, Anurag

A multiscale in silico approach to design polymer nanocomposites and scaffolds for bone tissue engineering applications is described in this study. This study focuses on the role of biomaterials design and selection, structural integrity and mechanical properties evolution during degradation and tissue regeneration in the successful design of polymer nanocomposite scaffolds. Polymer nanocomposite scaffolds are synthesized using aminoacid modified montmorillonite nanoclay with biomineralized hydroxyapatite and polycaprolactone (PCL/in situ HAPclay). Representative molecular models of polymer nanocomposite system are systematically developed using molecular dynamics (MD) technique and successfully validated using material characterization techniques. The constant force steered molecular dynamics (fSMD) simulation results indicate a two-phase nanomechanical behavior of the polymer nanocomposite. The MD and fSMD simulations results provide quantitative contributions of molecular interactions between different constituents of representative models and their effect on nanomechanical responses of nanoclay based polymer nanocomposite system. A finite element (FE) model of PCL/in situ HAPclay scaffold is built using micro-computed tomography images and bridging the nanomechanical properties obtained from fSMD simulations into the FE model. A new reduction factor, K is introduced into modeling results to consider the effect of wall porosity of the polymer scaffold. The effect of accelerated degradation under alkaline conditions and human osteoblast cells culture on the evolution of mechanical properties of scaffolds are studied and the damage mechanics based analytical models are developed. Finally, the novel multiscale models are developed that incorporate the complex molecular and microstructural properties, mechanical properties at nanoscale and structural levels and mechanical properties evolution during degradation and tissue formation in the polymer nanocomposite scaffold. Overall, this study provides a leap into methodologies for in silico design of biomaterials for bone tissue engineering applications. Furthermore, as a part of this work, a molecular dynamics study of rice DNA in the presence of single walled carbon nanotube is carried out to understand the role played by molecular interactions in the conformation changes of rice DNA. The simulations results showed wrapping of DNA onto SWCNT, breaking and forming of hydrogen bonds due to unzipping of Watson-Crick (WC) nucleobase pairs and forming of new non-WC nucleobase pairs in DNA.

Structure and Biophysics of CBFβ/RUNX and Its Translocation Products.

PubMed

Tahirov, Tahir H; Bushweller, John

2017-01-01

The core binding factor (CBF) transcription factor is somewhat unique in that it is composed of a DNA binding RUNX subunit (RUNX1, 2, or 3) and a non-DNA binding CBFβ subunit, which modulates RUNX protein activity by modulating the auto-inhibition of the RUNX subunits. Since the discovery of this fascinating transcription factor more than 20 years ago, there has been a robust effort to characterize the structure as well as the biochemical properties of CBF. More recently, these efforts have also extended to the fusion proteins that arise from the subunits of CBF in leukemia. This chapter highlights the work of numerous labs which has provided a detailed understanding of the structure and function of this transcription factor and its fusion proteins.

Simulation studies of DNA at the nanoscale: Interactions with proteins, polycations, and surfaces

NASA Astrophysics Data System (ADS)

Elder, Robert M.

Understanding the nanoscale interactions of DNA, a multifunctional biopolymer with sequence-dependent properties, with other biological and synthetic substrates and molecules is essential to advancing these technologies. This doctoral thesis research is aimed at understanding the thermodynamics and molecular-level structure when DNA interacts with proteins, polycations, and functionalized surfaces. First, we investigate the ability of a DNA damage recognition protein (HMGB1a) to bind to anti-cancer drug-induced DNA damage, seeking to explain how HMGB1a differentiates between the drugs in vivo. Using atomistic molecular dynamics simulations, we show that the structure of the drug-DNA molecule exhibits drug- and base sequence-dependence that explains some of the experimentally observed differential recognition of the drugs in various sequence contexts. Then, we show how steric hindrance from the drug decreases the deformability of the drug-DNA molecule, which decreases recognition by the protein, a concept that can be applied to rational drug design. Second, we study how polycation architecture and chemistry affect polycation-DNA binding so as to design optimal polycations for high efficiency gene (DNA) delivery. Using a multiscale computational approach involving atomistic and coarse-grained simulations, we examine how rearranging polylysine from a linear to a grafted architecture, and several aspects of the grafted architecture, affect polycation-DNA binding and the structure of polycation-DNA complexes. Next, going beyond lysine we examine how oligopeptide chemistry and sequence in the grafted architecture affects polycation-DNA binding and find that strategic placement of hydrophobic peptides might be used to tailor binding strength. Third, we study the adsorption and conformations of single-stranded DNA (an amphiphilic biopolymer) on model hydrophilic and hydrophobic surfaces. Short ssDNA oligomers adsorb to both surfaces with similar strength, with the strength of adsorption to the hydrophobic surface depending on the composition of the DNA strands, i.e. purine or pyrimidine bases. Additionally, DNA-surface and DNA-water interactions near the surfaces govern the adsorption. For longer ssDNA oligomers, the effects of surface chemistry and temperature on ssDNA conformations are rather small, but either the hydrophilic surface or increased temperature favor slightly more compact conformations due to energetic and entropic effects, respectively.

Selective detection of Mg2+ ions via enhanced fluorescence emission using Au–DNA nanocomposites

PubMed Central

Basu, Tanushree; Rana, Khyati; Das, Niranjan

2017-01-01

The biophysical properties of DNA-modified Au nanoparticles (AuNPs) have attracted a great deal of research interest for various applications in biosensing. AuNPs have strong binding capability to the phosphate and sugar groups in DNA, rendering unique physicochemical properties for detection of metal ions. The formation of Au–DNA nanocomposites is evident from the observed changes in the optical absorption, plasmon band, zeta potential, DLS particle size distribution, as well as TEM and AFM surface morphology analysis. Circular dichroism studies also revealed that DNA-functionalized AuNP binding caused a conformational change in the DNA structure. Due to the size and shape dependent plasmonic interactions of AuNPs (33–78 nm) with DNA, the resultant Au–DNA nanocomposites (NCs) exhibit superior fluorescence emission due to chemical binding with Ca2+, Fe2+ and Mg2+ ions. A significant increase in fluorescence emission (λex = 260 nm) of Au–DNA NCs was observed after selectively binding with Mg2+ ions (20–800 ppm) in an aqueous solution where a minimum of 100 ppm Mg2+ ions was detected based on the linearity of concentration versus fluorescence intensity curve (λem = 400 nm). The effectiveness of Au–DNA nanocomposites was further verified by comparing the known concentration (50–120 ppm) of Mg2+ ions in synthetic tap water and a real life sample of Gelusil (300–360 ppm Mg2+), a widely used antacid medicine. Therefore, this method could be a sensitive tool for the estimation of water hardness after careful preparation of a suitably designed Au–DNA nanostructure. PMID:28487819

DNA-guided nanoparticle assemblies

DOEpatents

Gang, Oleg; Nykypanchuk, Dmytro; Maye, Mathew; van der Lelie, Daniel

2013-07-16

In some embodiments, DNA-capped nanoparticles are used to define a degree of crystalline order in assemblies thereof. In some embodiments, thermodynamically reversible and stable body-centered cubic (bcc) structures, with particles occupying <.about.10% of the unit cell, are formed. Designs and pathways amenable to the crystallization of particle assemblies are identified. In some embodiments, a plasmonic crystal is provided. In some aspects, a method for controlling the properties of particle assemblages is provided. In some embodiments a catalyst is formed from nanoparticles linked by nucleic acid sequences and forming an open crystal structure with catalytically active agents attached to the crystal on its surface or in interstices.

Sanguinarine interacts with chromatin, modulates epigenetic modifications, and transcription in the context of chromatin.

PubMed

Selvi B, Ruthrotha; Pradhan, Suman Kalyan; Shandilya, Jayasha; Das, Chandrima; Sailaja, Badi Sri; Shankar G, Naga; Gadad, Shrikanth S; Reddy, Ashok; Dasgupta, Dipak; Kundu, Tapas K

2009-02-27

DNA-binding anticancer agents cause alteration in chromatin structure and dynamics. We report the dynamic interaction of the DNA intercalator and potential anticancer plant alkaloid, sanguinarine (SGR), with chromatin. Association of SGR with different levels of chromatin structure was enthalpy driven with micromolar dissociation constant. Apart from DNA, it binds with comparable affinity with core histones and induces chromatin aggregation. The dual binding property of SGR leads to inhibition of core histone modifications. Although it potently inhibits H3K9 methylation by G9a in vitro, H3K4 and H3R17 methylation are more profoundly inhibited in cells. SGR inhibits histone acetylation both in vitro and in vivo. It does not affect the in vitro transcription from DNA template but significantly represses acetylation-dependent chromatin transcription. SGR-mediated repression of epigenetic marks and the alteration of chromatin geography (nucleography) also result in the modulation of global gene expression. These data, conclusively, show an anticancer DNA binding intercalator as a modulator of chromatin modifications and transcription in the chromatin context.

Effect of calcium/sodium ion exchange on the osmotic properties and structure of polyelectrolyte gels.

PubMed

Horkay, Ferenc; Basser, Peter J; Hecht, Anne-Marie; Geissler, Erik

2015-12-01

We discuss the main findings of a long-term research program exploring the consequences of sodium/calcium ion exchange on the macroscopic osmotic and elastic properties, and the microscopic structure of representative synthetic polyelectrolyte (sodium polyacrylate, (polyacrylic acid)) and biopolymer gels (DNA). A common feature of these gels is that above a threshold calcium ion concentration, they exhibit a reversible volume phase transition. At the macroscopic level, the concentration dependence of the osmotic pressure shows that calcium ions influence primarily the third-order interaction term in the Flory-Huggins model of polymer solutions. Mechanical tests reveal that the elastic modulus is practically unaffected by the presence of calcium ions, indicating that ion bridging does not create permanent cross-links. At the microscopic level, small-angle neutron scattering shows that polyacrylic acid and DNA gels exhibit qualitatively similar structural features in spite of important differences (e.g. chain flexibility and chemical composition) between the two polymers. The main effect of calcium ions is that the neutron scattering intensity increases due to the decrease in the osmotic modulus. At the level of the counterion cloud around dissolved macroions, anomalous small-angle X-ray scattering measurements made on DNA indicate that divalent ions form a cylindrical sheath enveloping the chain, but they are not localized. Small-angle neutron scattering and small-angle X-ray scattering provide complementary information on the structure and interactions in polymer solutions and gels. © IMechE 2015.

Optical and structural properties of plasma-treated Cordyceps bassiana spores as studied by circular dichroism, absorption, and fluorescence spectroscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Geon Joon, E-mail: gjlee@kw.ac.kr; Sim, Geon Bo; Choi, Eun Ha

To understand the killing mechanism of fungal spores by plasma treatment, the optical, structural, and biological properties of the insect pathogenic fungus Cordyceps bassiana spores were studied. A nonthermal atmospheric-pressure plasma jet (APPJ) was used to treat the spores in aqueous solution. Optical emission spectra of the APPJ acquired in air indicated emission peaks corresponding to hydroxyl radicals and atomic oxygen. When the APPJ entered the aqueous solution, additional reactive species were derived from the interaction of plasma radicals with the aqueous solution. Fluorescence and absorption spectroscopy confirmed the generation of hydroxyl radicals and hydrogen peroxide in the plasma-activated watermore » (PAW). Spore counting showed that plasma treatment significantly reduced spore viability. Absorption spectroscopy, circular dichroism (CD) spectroscopy, and agarose gel electrophoresis of the DNA extracted from plasma-treated spores showed a reduction in spore DNA content. The magnitude of the dip in the CD spectrum was lower in the plasma-treated spores than in the control, indicating that plasma treatment causes structural modifications and/or damage to cellular components. Tryptophan fluorescence intensity was lower in the plasma-treated spores than in the control, suggesting that plasma treatment modified cell wall proteins. Changes in spore viability and DNA content were attributed to structural modification of the cell wall by reactive species coming from the APPJ and the PAW. Our results provided evidence that the plasma radicals and the derived reactive species play critical roles in fungal spore inactivation.« less

Optical and structural properties of plasma-treated Cordyceps bassiana spores as studied by circular dichroism, absorption, and fluorescence spectroscopy

NASA Astrophysics Data System (ADS)

Lee, Geon Joon; Sim, Geon Bo; Choi, Eun Ha; Kwon, Young-Wan; Kim, Jun Young; Jang, Siun; Kim, Seong Hwan

2015-01-01

To understand the killing mechanism of fungal spores by plasma treatment, the optical, structural, and biological properties of the insect pathogenic fungus Cordyceps bassiana spores were studied. A nonthermal atmospheric-pressure plasma jet (APPJ) was used to treat the spores in aqueous solution. Optical emission spectra of the APPJ acquired in air indicated emission peaks corresponding to hydroxyl radicals and atomic oxygen. When the APPJ entered the aqueous solution, additional reactive species were derived from the interaction of plasma radicals with the aqueous solution. Fluorescence and absorption spectroscopy confirmed the generation of hydroxyl radicals and hydrogen peroxide in the plasma-activated water (PAW). Spore counting showed that plasma treatment significantly reduced spore viability. Absorption spectroscopy, circular dichroism (CD) spectroscopy, and agarose gel electrophoresis of the DNA extracted from plasma-treated spores showed a reduction in spore DNA content. The magnitude of the dip in the CD spectrum was lower in the plasma-treated spores than in the control, indicating that plasma treatment causes structural modifications and/or damage to cellular components. Tryptophan fluorescence intensity was lower in the plasma-treated spores than in the control, suggesting that plasma treatment modified cell wall proteins. Changes in spore viability and DNA content were attributed to structural modification of the cell wall by reactive species coming from the APPJ and the PAW. Our results provided evidence that the plasma radicals and the derived reactive species play critical roles in fungal spore inactivation.

The structural changes of T7 RNA polymerase from transcription initiation to elongation

PubMed Central

Steitz, Thomas A

2010-01-01

Summary The structures of T7 RNA polymerase (T7 RNAP) captured in the initiation and elongation phases of transcription, as well as an intermediate stage provide insights into how this RNA polymerase protein can initiate RNA synthesis and synthesize 7 to 10 nucleotides of RNA while remaining bound to the DNA promoter site. Recently, the structures of T7 RNAP bound to it promoter DNA along with either a 7 nucleotide or 8 nucleotide transcript show an elongated product site resulting from a 40° or 45° rotation of the promoter and domain that binds it. The different functional properties of the initiation and elongation phases of transcription are illuminated from structures of the initiation and elongation complexes. Structural insights into the translocation of the product transcript of RNAP, its separation of the downstream duplex DNA and its removal of the transcript from the heteroduplex are provided by the structures of several states of nucleotide incorporation. A conformational change in the “fingers” domain that results from the binding or dissociation of incoming NTP or PPi appears to be associated with the state of translocation of T7 RNAP. PMID:19811903

A Case Study into Microbial Genome Assembly Gap Sequences and Finishing Strategies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Utturkar, Sagar M.; Klingeman, Dawn M.; Hurt, Jr., Richard A.

This study characterized regions of DNA which remained unassembled by either PacBio and Illumina sequencing technologies for seven bacterial genomes. Two genomes were manually finished using bioinformatics and PCR/Sanger sequencing approaches and regions not assembled by automated software were analyzed. Gaps present within Illumina assemblies mostly correspond to repetitive DNA regions such as multiple rRNA operon sequences. PacBio gap sequences were evaluated for several properties such as GC content, read coverage, gap length, ability to form strong secondary structures, and corresponding annotations. Our hypothesis that strong secondary DNA structures blocked DNA polymerases and contributed to gap sequences was not accepted.more » PacBio assemblies had few limitations overall and gaps were explained as cumulative effect of lower than average sequence coverage and repetitive sequences at contig termini. An important aspect of the present study is the compilation of biological features that interfered with assembly and included active transposons, multiple plasmid sequences, phage DNA integration, and large sequence duplication. Furthermore, our targeted genome finishing approach and systematic evaluation of the unassembled DNA will be useful for others looking to close, finish, and polish microbial genome sequences.« less

The implication of DNA bending energy for nucleosome positioning and sliding.

PubMed

Liu, Guoqing; Xing, Yongqiang; Zhao, Hongyu; Cai, Lu; Wang, Jianying

2018-06-11

Nucleosome not only directly affects cellular processes, such as DNA replication, recombination, and transcription, but also severs as a fundamentally important target of epigenetic modifications. Our previous study indicated that the bending property of DNA is important in nucleosome formation, particularly in predicting the dyad positions of nucleosomes on a DNA segment. Here, we investigated the role of bending energy in nucleosome positioning and sliding in depth to decipher sequence-directed mechanism. The results show that bending energy is a good physical index to predict the free energy in the process of nucleosome reconstitution in vitro. Our data also imply that there are at least 20% of the nucleosomes in budding yeast do not adopt canonical positioning, in which underlying sequences wrapped around histones are structurally symmetric. We also revealed distinct patterns of bending energy profile for distinctly organized chromatin structures, such as well-positioned nucleosomes, fuzzy nucleosomes, and linker regions and discussed nucleosome sliding in terms of bending energy. We proposed that the stability of a nucleosome is positively correlated with the strength of the bending anisotropy of DNA segment, and both accessibility and directionality of nucleosome sliding is likely to be modulated by diverse patterns of DNA bending energy profile.

A Case Study into Microbial Genome Assembly Gap Sequences and Finishing Strategies

DOE PAGES

Utturkar, Sagar M.; Klingeman, Dawn M.; Hurt, Jr., Richard A.; ...

2017-07-18

This study characterized regions of DNA which remained unassembled by either PacBio and Illumina sequencing technologies for seven bacterial genomes. Two genomes were manually finished using bioinformatics and PCR/Sanger sequencing approaches and regions not assembled by automated software were analyzed. Gaps present within Illumina assemblies mostly correspond to repetitive DNA regions such as multiple rRNA operon sequences. PacBio gap sequences were evaluated for several properties such as GC content, read coverage, gap length, ability to form strong secondary structures, and corresponding annotations. Our hypothesis that strong secondary DNA structures blocked DNA polymerases and contributed to gap sequences was not accepted.more » PacBio assemblies had few limitations overall and gaps were explained as cumulative effect of lower than average sequence coverage and repetitive sequences at contig termini. An important aspect of the present study is the compilation of biological features that interfered with assembly and included active transposons, multiple plasmid sequences, phage DNA integration, and large sequence duplication. Furthermore, our targeted genome finishing approach and systematic evaluation of the unassembled DNA will be useful for others looking to close, finish, and polish microbial genome sequences.« less

A Case Study into Microbial Genome Assembly Gap Sequences and Finishing Strategies

PubMed Central

Utturkar, Sagar M.; Klingeman, Dawn M.; Hurt, Richard A.; Brown, Steven D.

2017-01-01

This study characterized regions of DNA which remained unassembled by either PacBio and Illumina sequencing technologies for seven bacterial genomes. Two genomes were manually finished using bioinformatics and PCR/Sanger sequencing approaches and regions not assembled by automated software were analyzed. Gaps present within Illumina assemblies mostly correspond to repetitive DNA regions such as multiple rRNA operon sequences. PacBio gap sequences were evaluated for several properties such as GC content, read coverage, gap length, ability to form strong secondary structures, and corresponding annotations. Our hypothesis that strong secondary DNA structures blocked DNA polymerases and contributed to gap sequences was not accepted. PacBio assemblies had few limitations overall and gaps were explained as cumulative effect of lower than average sequence coverage and repetitive sequences at contig termini. An important aspect of the present study is the compilation of biological features that interfered with assembly and included active transposons, multiple plasmid sequences, phage DNA integration, and large sequence duplication. Our targeted genome finishing approach and systematic evaluation of the unassembled DNA will be useful for others looking to close, finish, and polish microbial genome sequences. PMID:28769883

Structural and biophysical analysis of interactions between cod and human uracil-DNA N-glycosylase (UNG) and UNG inhibitor (Ugi)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Assefa, Netsanet Gizaw; Niiranen, Laila; University of Turku, FIN-20014 Turku

2014-08-01

A structural and biophysical study of the interactions between cod and human uracil-DNA N-glycosylase (UNG) and their inhibitor Ugi is presented. The stronger interaction between cod UNG and Ugi can be explained by a greater positive electrostatic surface potential. Uracil-DNA N-glycosylase from Atlantic cod (cUNG) shows cold-adapted features such as high catalytic efficiency, a low temperature optimum for activity and reduced thermal stability compared with its mesophilic homologue human UNG (hUNG). In order to understand the role of the enzyme–substrate interaction related to the cold-adapted properties, the structure of cUNG in complex with a bacteriophage encoded natural UNG inhibitor (Ugi)more » has been determined. The interaction has also been analyzed by isothermal titration calorimetry (ITC). The crystal structure of cUNG–Ugi was determined to a resolution of 1.9 Å with eight complexes in the asymmetric unit related through noncrystallographic symmetry. A comparison of the cUNG–Ugi complex with previously determined structures of UNG–Ugi shows that they are very similar, and confirmed the nucleotide-mimicking properties of Ugi. Biophysically, the interaction between cUNG and Ugi is very strong and shows a binding constant (K{sub b}) which is one order of magnitude larger than that for hUNG–Ugi. The binding of both cUNG and hUNG to Ugi was shown to be favoured by both enthalpic and entropic forces; however, the binding of cUNG to Ugi is mainly dominated by enthalpy, while the entropic term is dominant for hUNG. The observed differences in the binding properties may be explained by an overall greater positive electrostatic surface potential in the protein–Ugi interface of cUNG and the slightly more hydrophobic surface of hUNG.« less

RNA-DNA Triplex Formation by Long Noncoding RNAs.

PubMed

Li, Yue; Syed, Junetha; Sugiyama, Hiroshi

2016-11-17

Long noncoding RNAs (lncRNAs) play a pivotal role in the regulation of biological processes through various mechanisms that are not fully understood. Proposed mechanisms include regulation based on RNA-protein interactions, as well as RNA-RNA interactions and RNA-DNA interactions. Here, we focus on one possible mechanism that lncRNA might be using to impact biological function, the RNA-DNA triplex formation. We summarize currently available examples of lncRNA triplex formation and discuss the details surrounding orientation of triplex formation as one of the key properties guiding this process. We propose that symmetrical triplex-forming motifs, especially those in cis-acting lncRNAs, favor triplex formation. We also consider the effects of lncRNA structures, protein or ligand binding, and chromatin structures on the lncRNAs triplex formation. Copyright © 2016 Elsevier Ltd. All rights reserved.

Excluded volume and ion-ion correlation effects on the ionic atmosphere around B-DNA: Theory, simulations, and experiments

PubMed Central

Ovanesyan, Zaven; Fenley, Marcia O.; Guerrero-García, Guillermo Iván; Olvera de la Cruz, Mónica

2014-01-01

The ionic atmosphere around a nucleic acid regulates its stability in aqueous salt solutions. One major source of complexity in biological activities involving nucleic acids arises from the strong influence of the surrounding ions and water molecules on their structural and thermodynamic properties. Here, we implement a classical density functional theory for cylindrical polyelectrolytes embedded in aqueous electrolytes containing explicit (neutral hard sphere) water molecules at experimental solvent concentrations. Our approach allows us to include ion correlations as well as solvent and ion excluded volume effects for studying the structural and thermodynamic properties of highly charged cylindrical polyelectrolytes. Several models of size and charge asymmetric mixtures of aqueous electrolytes at physiological concentrations are studied. Our results are in good agreement with Monte Carlo simulations. Our numerical calculations display significant differences in the ion density profiles for the different aqueous electrolyte models studied. However, similar results regarding the excess number of ions adsorbed to the B-DNA molecule are predicted by our theoretical approach for different aqueous electrolyte models. These findings suggest that ion counting experimental data should not be used alone to validate the performance of aqueous DNA-electrolyte models. PMID:25494770

Studying the orientation of bio-objects by nematic liquid crystals

NASA Astrophysics Data System (ADS)

Zubtsova, Yu. A.; Kamanin, A. A.; Kamanina, N. V.

2017-05-01

We have studied the ability of a liquid-crystal (LC) matrix to visualize and orient DNA molecules. It is established that the relief of the interface between the LC mesophase and conducting contact can be improved without using an additional high-ohmic polymer layer. Spectroscopic and ellipsometric techniques revealed changes in the refractive properties and structure of composites. The obtained results can be used in creating devices for rapid DNA testing with retained form of biostructures.

Direct electrical and mechanical characterization of in situ generated DNA between the tips of silicon nanotweezers (SNT).

PubMed

Karsten, Stanislav L; Kumemura, Momoko; Jalabert, Laurent; Lafitte, Nicolas; Kudo, Lili C; Collard, Dominique; Fujita, Hiroyuki

2016-05-24

Previously, we reported the application of micromachined silicon nanotweezers (SNT) integrated with a comb-drive actuator and capacitive sensors for capturing and mechanical characterization of DNA bundles. Here, we demonstrate direct DNA amplification on such a MEMS structure with subsequent electrical and mechanical characterization of a single stranded DNA (ssDNA) bundle generated between the tips of SNT via isothermal rolling circle amplification (RCA) and dielectrophoresis (DEP). An in situ generated ssDNA bundle was visualized and evaluated via electrical conductivity (I-V) and mechanical frequency response measurements. Colloidal gold nanoparticles significantly enhanced (P < 0.01) the electrical properties of thin ssDNA bundles. The proposed technology allows direct in situ synthesis of DNA with a predefined sequence on the tips of a MEMS sensor device, such as SNT, followed by direct DNA electrical and mechanical characterization. In addition, our data provides a "proof-of-principle" for the feasibility of the on-chip label free DNA detection device that can be used for a variety of biomedical applications focused on sequence specific DNA detection.

Effects of Near Infrared Radiation on DNA. DLS and ATR-FTIR Study

NASA Astrophysics Data System (ADS)

Szymborska-Małek, Katarzyna; Komorowska, Małgorzata; Gąsior-Głogowska, Marlena

2018-01-01

We presume that the primary effect of Near Infrared (NIR) radiation on aqueous solutions of biological molecules concerns modification of hydrogen bonded structures mainly the global and the hydration shell water molecules. Since water has a significant influence on the DNA structure, we expect that the thermal stability of DNA could be modified by NIR radiation. The herring sperm DNA was exposed to NIR radiation (700-1100 nm) for 5, 10, and 20 min periods. The temperature dependent infrared measurements were done for the thin films formed on the diamond ATR crystal from evaporated DNA solutions exposed and unexposed to NIR radiation. For the NIR-treated samples (at room temperature) the B form was better conserved than in the control sample independently of the irradiation period. Above 50 °C a considerable increase in the A form was only observed for 10 min NIR exposed samples. The hydrodynamic radius, (Rh), studied by the dynamic light scattering, showed drastic decrease with the increasing irradiation time. Principal components analysis (PCA) allowed to detect the spectral features correlated with the NIR effect and thermal stability of the DNA films. Obtained results strongly support the idea that the photoionization of water by NIR radiation in presence of DNA molecules is the main factor influencing on its physicochemical properties.

A comprehensive approach to ascertain the binding mode of curcumin with DNA

NASA Astrophysics Data System (ADS)

Haris, P.; Mary, Varughese; Aparna, P.; Dileep, K. V.; Sudarsanakumar, C.

2017-03-01

Curcumin is a natural phytochemical from the rhizoma of Curcuma longa, the popular Indian spice that exhibits a wide range of pharmacological properties like antioxidant, anticancer, anti-inflammatory, antitumor, and antiviral activities. In the published literatures we can see different studies and arguments on the interaction of curcumin with DNA. The intercalative binding, groove binding and no binding of curcumin with DNA were reported. In this context, we conducted a detailed study to understand the mechanism of recognition of dimethylsulfoxide-solubilized curcumin by DNA. The interaction of curcumin with calf thymus DNA (ctDNA) was confirmed by agarose gel electrophoresis. The nature of binding and energetics of interaction were studied by Isothermal Titration Calorimetry (ITC), Differential Scanning Calorimetry (DSC), UV-visible, fluorescence and melting temperature (Tm) analysis. The experimental data were compared with molecular modeling studies. Our investigation confirmed that dimethylsulfoxide-solubilized curcumin binds in the minor groove of the ctDNA without causing significant structural alteration to the DNA.

Deciphering the mechanism of interaction of edifenphos with calf thymus DNA

NASA Astrophysics Data System (ADS)

Ahmad, Ajaz; Ahmad, Masood

2018-01-01

Edifenphos is an important organophosphate pesticide with many antifungal and anti-insecticidal properties but it may cause potential hazards to human health. In this work, we have tried to explore the binding mode of action and mechanism of edifenphos to calf thymus DNA (CT-DNA). Several experiments such as ultraviolet-visible absorption spectra and emission spectroscopy showed complex formation between edifenphos and CT-DNA and low binding constant values supporting groove binding mode. These results were further confirmed by circular dichroism (CD), CT-DNA melting studies, viscosity measurements, density functional theory and molecular docking. CD study suggests that edifenphos does not alter native structure of CT-DNA. Isothermal calorimetry reveals that binding of edifenphos with CT-DNA is enthalpy driven process. Competitive binding assay and effect of ionic strength showed that edifenphos binds to CT-DNA via groove binding manner. Hence, edifenphos is a minor groove binder preferably interacting with A-T regions with docking score - 6.84 kJ/mol.

Amino Acid Interaction (INTAA) web server.

PubMed

Galgonek, Jakub; Vymetal, Jirí; Jakubec, David; Vondrášek, Jirí

2017-07-03

Large biomolecules-proteins and nucleic acids-are composed of building blocks which define their identity, properties and binding capabilities. In order to shed light on the energetic side of interactions of amino acids between themselves and with deoxyribonucleotides, we present the Amino Acid Interaction web server (http://bioinfo.uochb.cas.cz/INTAA/). INTAA offers the calculation of the residue Interaction Energy Matrix for any protein structure (deposited in Protein Data Bank or submitted by the user) and a comprehensive analysis of the interfaces in protein-DNA complexes. The Interaction Energy Matrix web application aims to identify key residues within protein structures which contribute significantly to the stability of the protein. The application provides an interactive user interface enhanced by 3D structure viewer for efficient visualization of pairwise and net interaction energies of individual amino acids, side chains and backbones. The protein-DNA interaction analysis part of the web server allows the user to view the relative abundance of various configurations of amino acid-deoxyribonucleotide pairs found at the protein-DNA interface and the interaction energies corresponding to these configurations calculated using a molecular mechanical force field. The effects of the sugar-phosphate moiety and of the dielectric properties of the solvent on the interaction energies can be studied for the various configurations. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Structural modelling and molecular dynamics of a multi-stress responsive WRKY TF-DNA complex towards elucidating its role in stress signalling mechanisms in chickpea.

PubMed

Konda, Aravind Kumar; Farmer, Rohit; Soren, Khela Ram; P S, Shanmugavadivel; Setti, Aravind

2017-07-28

Chickpea is a premier food legume crop with high nutritional quality and attains prime importance in the current era of 795 million people being undernourished worldwide. Chickpea production encounters setbacks due to various stresses and understanding the role of key transcription factors (TFs) involved in multiple stresses becomes inevitable. We have recently identified a multi-stress responsive WRKY TF in chickpea. The present study was conducted to predict the structure of WRKY TF to identify the DNA-interacting residues and decipher DNA-protein interactions. Comparative modelling approach produced 3D model of the WRKY TF with good stereochemistry, local/global quality and further revealed W19, R20, K21, and Y22 motifs within a vicinity of 5 Å to the DNA amongst R18, G23, Q24, K25, Y36, Y37, R38 and K47 and these positions were equivalent to the 2LEX WRKY domain of Arabidopsis. Molecular simulations analysis of reference protein -PDB ID 2LEX, along with Car-WRKY TF modelled structure with the DNA coordinates derived from PDB ID 2LEX and docked using HADDOCK were executed. Root Mean Square (RMS) Deviation and RMS Fluctuation values yielded consistently stable trajectories over 50 ns simulation. Strengthening the obtained results, neither radius of gyration, distance and total energy showed any signs of DNA-WRKY complex falling apart nor any significant dissociation event over 50 ns run. Therefore, the study provides first insights into the structural properties of multi-stress responsive WRKY TF-DNA complex in chickpea, enabling genome wide identification of TF binding sites and thereby deciphers their gene regulatory networks.

Electronic couplings and on-site energies for hole transfer in DNA: Systematic quantum mechanical/molecular dynamic study

NASA Astrophysics Data System (ADS)

Voityuk, Alexander A.

2008-03-01

The electron hole transfer (HT) properties of DNA are substantially affected by thermal fluctuations of the π stack structure. Depending on the mutual position of neighboring nucleobases, electronic coupling V may change by several orders of magnitude. In the present paper, we report the results of systematic QM/molecular dynamic (MD) calculations of the electronic couplings and on-site energies for the hole transfer. Based on 15ns MD trajectories for several DNA oligomers, we calculate the average coupling squares ⟨V2⟩ and the energies of basepair triplets XG +Y and XA +Y, where X, Y =G, A, T, and C. For each of the 32 systems, 15 000 conformations separated by 1ps are considered. The three-state generalized Mulliken-Hush method is used to derive electronic couplings for HT between neighboring basepairs. The adiabatic energies and dipole moment matrix elements are computed within the INDO/S method. We compare the rms values of V with the couplings estimated for the idealized B-DNA structure and show that in several important cases the couplings calculated for the idealized B-DNA structure are considerably underestimated. The rms values for intrastrand couplings G-G, A-A, G-A, and A-G are found to be similar, ˜0.07eV, while the interstrand couplings are quite different. The energies of hole states G+ and A+ in the stack depend on the nature of the neighboring pairs. The XG +Y are by 0.5eV more stable than XA +Y. The thermal fluctuations of the DNA structure facilitate the HT process from guanine to adenine. The tabulated couplings and on-site energies can be used as reference parameters in theoretical and computational studies of HT processes in DNA.

Transcription factor-based biosensors enlightened by the analyte

PubMed Central

Fernandez-López, Raul; Ruiz, Raul; de la Cruz, Fernando; Moncalián, Gabriel

2015-01-01

Whole cell biosensors (WCBs) have multiple applications for environmental monitoring, detecting a wide range of pollutants. WCBs depend critically on the sensitivity and specificity of the transcription factor (TF) used to detect the analyte. We describe the mechanism of regulation and the structural and biochemical properties of TF families that are used, or could be used, for the development of environmental WCBs. Focusing on the chemical nature of the analyte, we review TFs that respond to aromatic compounds (XylS-AraC, XylR-NtrC, and LysR), metal ions (MerR, ArsR, DtxR, Fur, and NikR) or antibiotics (TetR and MarR). Analyzing the structural domains involved in DNA recognition, we highlight the similitudes in the DNA binding domains (DBDs) of these TF families. Opposite to DBDs, the wide range of analytes detected by TFs results in a diversity of structures at the effector binding domain. The modular architecture of TFs opens the possibility of engineering TFs with hybrid DNA and effector specificities. Yet, the lack of a crisp correlation between structural domains and specific functions makes this a challenging task. PMID:26191047

Transcription factor-based biosensors enlightened by the analyte.

PubMed

Fernandez-López, Raul; Ruiz, Raul; de la Cruz, Fernando; Moncalián, Gabriel

2015-01-01

Whole cell biosensors (WCBs) have multiple applications for environmental monitoring, detecting a wide range of pollutants. WCBs depend critically on the sensitivity and specificity of the transcription factor (TF) used to detect the analyte. We describe the mechanism of regulation and the structural and biochemical properties of TF families that are used, or could be used, for the development of environmental WCBs. Focusing on the chemical nature of the analyte, we review TFs that respond to aromatic compounds (XylS-AraC, XylR-NtrC, and LysR), metal ions (MerR, ArsR, DtxR, Fur, and NikR) or antibiotics (TetR and MarR). Analyzing the structural domains involved in DNA recognition, we highlight the similitudes in the DNA binding domains (DBDs) of these TF families. Opposite to DBDs, the wide range of analytes detected by TFs results in a diversity of structures at the effector binding domain. The modular architecture of TFs opens the possibility of engineering TFs with hybrid DNA and effector specificities. Yet, the lack of a crisp correlation between structural domains and specific functions makes this a challenging task.

Dynamic DNA nanotechnology using strand-displacement reactions

NASA Astrophysics Data System (ADS)

Zhang, David Yu; Seelig, Georg

2011-02-01

The specificity and predictability of Watson-Crick base pairing make DNA a powerful and versatile material for engineering at the nanoscale. This has enabled the construction of a diverse and rapidly growing set of DNA nanostructures and nanodevices through the programmed hybridization of complementary strands. Although it had initially focused on the self-assembly of static structures, DNA nanotechnology is now also becoming increasingly attractive for engineering systems with interesting dynamic properties. Various devices, including circuits, catalytic amplifiers, autonomous molecular motors and reconfigurable nanostructures, have recently been rationally designed to use DNA strand-displacement reactions, in which two strands with partial or full complementarity hybridize, displacing in the process one or more pre-hybridized strands. This mechanism allows for the kinetic control of reaction pathways. Here, we review DNA strand-displacement-based devices, and look at how this relatively simple mechanism can lead to a surprising diversity of dynamic behaviour.

Control of DNA-Functionalized Nanoparticle Assembly

NASA Astrophysics Data System (ADS)

Olvera de La Cruz, Monica

Directed crystallization of a large variety of nanoparticles, including proteins, via DNA hybridization kinetics has led to unique materials with a broad range of crystal symmetries. The nanoparticles are functionalized with DNA chains that link neighboring functionalized units. The shape of the nanoparticle, the DNA length, the sequence of the hybridizing DNA linker and the grafting density determine the crystal symmetries and lattice spacing. By carefully selecting these parameters one can, in principle, achieve all the symmetries found for both atomic and colloidal crystals of asymmetric shapes as well as new symmetries, and drive transitions between them. A scale-accurate coarse-grained model with explicit DNA chains provides the design parameters, including degree of hybridization, to achieve specific crystal structures. The model also provides surface energy values to determine the shape of defect-free single crystals with macroscopic anisotropic properties, as well as the parameters to develop colloidal models that reproduce both the shape of single crystals and their growth kinetics.

Conformational gating of DNA conductance

PubMed Central

Artés, Juan Manuel; Li, Yuanhui; Qi, Jianqing; Anantram, M. P.; Hihath, Joshua

2015-01-01

DNA is a promising molecule for applications in molecular electronics because of its unique electronic and self-assembly properties. Here we report that the conductance of DNA duplexes increases by approximately one order of magnitude when its conformation is changed from the B-form to the A-form. This large conductance increase is fully reversible, and by controlling the chemical environment, the conductance can be repeatedly switched between the two values. The conductance of the two conformations displays weak length dependencies, as is expected for guanine-rich sequences, and can be fit with a coherence-corrected hopping model. These results are supported by ab initio electronic structure calculations that indicate that the highest occupied molecular orbital is more disperse in the A-form DNA case. These results demonstrate that DNA can behave as a promising molecular switch for molecular electronics applications and also provide additional insights into the huge dispersion of DNA conductance values found in the literature. PMID:26648400

Conformational gating of DNA conductance.

PubMed

Artés, Juan Manuel; Li, Yuanhui; Qi, Jianqing; Anantram, M P; Hihath, Joshua

2015-12-09

DNA is a promising molecule for applications in molecular electronics because of its unique electronic and self-assembly properties. Here we report that the conductance of DNA duplexes increases by approximately one order of magnitude when its conformation is changed from the B-form to the A-form. This large conductance increase is fully reversible, and by controlling the chemical environment, the conductance can be repeatedly switched between the two values. The conductance of the two conformations displays weak length dependencies, as is expected for guanine-rich sequences, and can be fit with a coherence-corrected hopping model. These results are supported by ab initio electronic structure calculations that indicate that the highest occupied molecular orbital is more disperse in the A-form DNA case. These results demonstrate that DNA can behave as a promising molecular switch for molecular electronics applications and also provide additional insights into the huge dispersion of DNA conductance values found in the literature.

Cellular DNA breakage by soy isoflavone genistein and its methylated structural analogue biochanin A.

PubMed

Ullah, Mohd Fahad; Shamim, Uzma; Hanif, Sarmad; Azmi, Asfar S; Hadi, Sheikh M

2009-11-01

Epidemiological studies have indicated that populations with high isoflavone intake through soy consumption have lower rates of breast, prostate, and colon cancer. The isoflavone polyphenol genistein in soybean is considered to be a potent chemopreventive agent against cancer. In order to explore the chemical basis of chemopreventive activity of genistein, in this paper we have examined the structure-activity relationship between genistein and its structural analogue biochanin A. We show that both genistein and its methylated derivative biochanin A are able to mobilize nuclear copper in human lymphocyte, leading to degradation of cellular DNA. However, the relative rate of DNA breakage was greater in the case of genistein. Further, the cellular DNA degradation was inhibited by copper chelator (neocuproine/bathocuproine) but not by compounds that specifically bind iron and zinc (desferrioxamine mesylate and histidine, respectively). We also compared the antioxidant activity of the two isoflavones against tert-butylhydroperoxide-induced oxidative breakage in lymphocytes. Again genistein was found to be more effective than biochanin A in providing protection against oxidative stress induced by tert-butylhydroperoxide. It would therefore appear that the structural features of isoflavones that are important for antioxidant properties are also the ones that contribute to their pro-oxidant action through a mechanism that involves redox cycling of chromatin-bound nuclear copper.

Ultrasensitive label-free detection of DNA hybridization by sapphire-based graphene field-effect transistor biosensor

NASA Astrophysics Data System (ADS)

Xu, Shicai; Jiang, Shouzhen; Zhang, Chao; Yue, Weiwei; Zou, Yan; Wang, Guiying; Liu, Huilan; Zhang, Xiumei; Li, Mingzhen; Zhu, Zhanshou; Wang, Jihua

2018-01-01

Graphene has attracted much attention in biosensing applications for its unique properties. Because of one-atom layer structure, every atom of graphene is exposed to the environment, making the electronic properties of graphene are very sensitive to charged analytes. Therefore, graphene is an ideal material for transistors in high-performance sensors. Chemical vapor deposition (CVD) method has been demonstrated the most successful method for fabricating large area graphene. However, the conventional CVD methods can only grow graphene on metallic substrate and the graphene has to be transferred to the insulating substrate for further device fabrication. The transfer process creates wrinkles, cracks, or tears on the graphene, which severely degrade electrical properties of graphene. These factors severely degrade the sensing performance of graphene. Here, we directly fabricated graphene on sapphire substrate by high temperature CVD without the use of metal catalysts. The sapphire-based graphene was patterned and make into a DNA biosensor in the configuration of field-effect transistor. The sensors show high performance and achieve the DNA detection sensitivity as low as 100 fM (10-13 M), which is at least 10 times lower than prior transferred CVD G-FET DNA sensors. The use of the sapphire-based G-FETs suggests a promising future for biosensing applications.

Structural and thermodynamic analysis of modified nucleosides in self-assembled DNA cross-tiles.

PubMed

Hakker, Lauren; Marchi, Alexandria N; Harris, Kimberly A; LaBean, Thomas H; Agris, Paul F

2014-01-01

DNA Holliday junctions are important natural strand-exchange structures that form during homologous recombination. Immobile four-arm junctions, analogs to Holliday junctions, have been designed to self-assemble into cross-tile structures by maximizing Watson-Crick base pairing and fixed crossover points. The cross-tiles, self-assembled from base pair recognition between designed single-stranded DNAs, form higher order lattice structures through cohesion of self-associating sticky ends. These cross-tiles have 16 unpaired nucleosides in the central loop at the junction of the four duplex stems. The importance of the centralized unpaired nucleosides to the structure's thermodynamic stability and self-assembly is unknown. Cross-tile DNA nanostructures were designed and constructed from nine single-stranded DNAs with four shell strands, four arms, and a central loop containing 16 unpaired bases. The 16 unpaired bases were either 2'-deoxyribothymidines, 2'-O-methylribouridines, or abasic 1',2'-dideoxyribonucleosides. Thermodynamic profiles and structural base-stacking contributions were assessed using UV absorption spectroscopy during thermal denaturation and circular dichroism spectroscopy, respectively, and the resulting structures were observed by atomic force microscopy. There were surprisingly significant changes in the thermodynamic and structural properties of lattice formation as a result of altering only the 16 unpaired, centralized nucleosides. The 16 unpaired 2'-O-methyluridines were stabilizing and produced uniform tubular structures. In contrast, the abasic nucleosides were destabilizing producing a mixture of structures. These results strongly indicate the importance of a small number of centrally located unpaired nucleosides within the structures. Since minor modifications lead to palpable changes in lattice formation, DNA cross-tiles present an easily manipulated structure convenient for applications in biomedical and biosensing devices.

Reviving the RNA World: An Insight into the Appearance of RNA Methyltransferases

PubMed Central

Rana, Ajay K.; Ankri, Serge

2016-01-01

RNA, the earliest genetic and catalytic molecule, has a relatively delicate and labile chemical structure, when compared to DNA. It is prone to be damaged by alkali, heat, nucleases, or stress conditions. One mechanism to protect RNA or DNA from damage is through site-specific methylation. Here, we propose that RNA methylation began prior to DNA methylation in the early forms of life evolving on Earth. In this article, the biochemical properties of some RNA methyltransferases (MTases), such as 2′-O-MTases (Rlml/RlmN), spOUT MTases and the NSun2 MTases are dissected for the insight they provide on the transition from an RNA world to our present RNA/DNA/protein world. PMID:27375676

DNA-Based Self-Assembly of Fluorescent Nanodiamonds.

PubMed

Zhang, Tao; Neumann, Andre; Lindlau, Jessica; Wu, Yuzhou; Pramanik, Goutam; Naydenov, Boris; Jelezko, Fedor; Schüder, Florian; Huber, Sebastian; Huber, Marinus; Stehr, Florian; Högele, Alexander; Weil, Tanja; Liedl, Tim

2015-08-12

As a step toward deterministic and scalable assembly of ordered spin arrays we here demonstrate a bottom-up approach to position fluorescent nanodiamonds (NDs) with nanometer precision on DNA origami structures. We have realized a reliable and broadly applicable surface modification strategy that results in DNA-functionalized and perfectly dispersed NDs that were then self-assembled in predefined geometries. With optical studies we show that the fluorescence properties of the nitrogen-vacancy color centers in NDs are preserved during surface modification and DNA assembly. As this method allows the nanoscale arrangement of fluorescent NDs together with other optically active components in complex geometries, applications based on self-assembled spin lattices or plasmon-enhanced spin sensors as well as improved fluorescent labeling for bioimaging could be envisioned.

Investigating the Structural Impacts of I64T and P311S Mutations in APE1-DNA Complex: A Molecular Dynamics Approach

PubMed Central

Doss, C. George Priya; NagaSundaram, N.

2012-01-01

Background Elucidating the molecular dynamic behavior of Protein-DNA complex upon mutation is crucial in current genomics. Molecular dynamics approach reveals the changes on incorporation of variants that dictate the structure and function of Protein-DNA complexes. Deleterious mutations in APE1 protein modify the physicochemical property of amino acids that affect the protein stability and dynamic behavior. Further, these mutations disrupt the binding sites and prohibit the protein to form complexes with its interacting DNA. Principal Findings In this study, we developed a rapid and cost-effective method to analyze variants in APE1 gene that are associated with disease susceptibility and evaluated their impacts on APE1-DNA complex dynamic behavior. Initially, two different in silico approaches were used to identify deleterious variants in APE1 gene. Deleterious scores that overlap in these approaches were taken in concern and based on it, two nsSNPs with IDs rs61730854 (I64T) and rs1803120 (P311S) were taken further for structural analysis. Significance Different parameters such as RMSD, RMSF, salt bridge, H-bonds and SASA applied in Molecular dynamic study reveals that predicted deleterious variants I64T and P311S alters the structure as well as affect the stability of APE1-DNA interacting functions. This study addresses such new methods for validating functional polymorphisms of human APE1 which is critically involved in causing deficit in repair capacity, which in turn leads to genetic instability and carcinogenesis. PMID:22384055

Ionic Conductivity, Structural Deformation and Programmable Anisotropy of DNA Origami in Electric Field

PubMed Central

Li, Chen-Yu; Hemmig, Elisa A.; Kong, Jinglin; Yoo, Jejoong; Hernández-Ainsa, Silvia

2015-01-01

The DNA origami technique can enable functionalization of inorganic structures for single-molecule electric current recordings. Experiments have shown that several layers of DNA molecules—a DNA origami plate— placed on top of a solid-state nanopore is permeable to ions. Here, we report a comprehensive characterization of the ionic conductivity of DNA origami plates by means of all-atom molecular dynamics (MD) simulations and nanocapillary electric current recordings. Using the MD method, we characterize the ionic conductivity of several origami constructs, revealing the local distribution of ions, the distribution of the electrostatic potential and contribution of different molecular species to the current. The simulations determine the dependence of the ionic conductivity on the applied voltage, the number of DNA layers, the nucleotide content and the lattice type of the plates. We demonstrate that increasing the concentration of Mg2+ ions makes the origami plates more compact, reducing their conductivity. The conductance of a DNA origami plate on top of a solid-state nanopore is determined by the two competing effects: bending of the DNA origami plate that reduces the current and separation of the DNA origami layers that increases the current. The latter is produced by the electro-osmotic flow and is reversible at the time scale of a hundred nanoseconds. The conductance of a DNA origami object is found to depend on its orientation, reaching maximum when the electric field aligns with the direction of the DNA helices. Our work demonstrates feasibility of programming the electrical properties of a self-assembled nanoscale object using DNA. PMID:25623807

Ionic conductivity, structural deformation, and programmable anisotropy of DNA origami in electric field.

PubMed

Li, Chen-Yu; Hemmig, Elisa A; Kong, Jinglin; Yoo, Jejoong; Hernández-Ainsa, Silvia; Keyser, Ulrich F; Aksimentiev, Aleksei

2015-02-24

The DNA origami technique can enable functionalization of inorganic structures for single-molecule electric current recordings. Experiments have shown that several layers of DNA molecules, a DNA origami plate, placed on top of a solid-state nanopore is permeable to ions. Here, we report a comprehensive characterization of the ionic conductivity of DNA origami plates by means of all-atom molecular dynamics (MD) simulations and nanocapillary electric current recordings. Using the MD method, we characterize the ionic conductivity of several origami constructs, revealing the local distribution of ions, the distribution of the electrostatic potential and contribution of different molecular species to the current. The simulations determine the dependence of the ionic conductivity on the applied voltage, the number of DNA layers, the nucleotide content and the lattice type of the plates. We demonstrate that increasing the concentration of Mg(2+) ions makes the origami plates more compact, reducing their conductivity. The conductance of a DNA origami plate on top of a solid-state nanopore is determined by the two competing effects: bending of the DNA origami plate that reduces the current and separation of the DNA origami layers that increases the current. The latter is produced by the electro-osmotic flow and is reversible at the time scale of a hundred nanoseconds. The conductance of a DNA origami object is found to depend on its orientation, reaching maximum when the electric field aligns with the direction of the DNA helices. Our work demonstrates feasibility of programming the electrical properties of a self-assembled nanoscale object using DNA.

Uncovering the self-assembly of DNA nanostructures by thermodynamics and kinetics.

PubMed

Wei, Xixi; Nangreave, Jeanette; Liu, Yan

2014-06-17

CONSPECTUS: DNA nanotechnology is one of the most flourishing interdisciplinary research fields. DNA nanostructures can be designed to self-assemble into a variety of periodic or aperiodic patterns of different shapes and length scales. They can be used as scaffolds for organizing other nanoparticles, proteins, and chemical groups, leveraging their functions for creating complex bioinspired materials that may serve as smart drug delivery systems, in vitro or in vivo biomolecular computing platforms, and diagnostic devices. Achieving optimal structural features, efficient assembly protocols, and precise functional group positioning and modification requires a thorough understanding of the thermodynamics and kinetics of the DNA nanostructure self-assembly process. The most common real-time measurement strategies include monitoring changes in UV absorbance based on the hyperchromic effect of DNA, and the emission signal changes of DNA intercalating dyes or covalently conjugated fluorescent dyes/pairs that accompany temperature dependent structural changes. Thermodynamic studies of a variety of DNA nanostructures have been performed, from simple double stranded DNA formation to more complex origami assembly. The key parameters that have been evaluated in terms of stability and cooperativity include the overall dimensions, the folding path of the scaffold, crossover and nick point arrangement, length and sequence of single strands, and salt and ion concentrations. DNA tile-tile interactions through sticky end hybridization have also been analyzed, and the steric inhibition and rigidity of tiles turn out to be important factors. Many kinetic studies have also been reported, and most are based on double stranded DNA formation. A two-state assumption and the hypothesis of several intermediate states have been applied to determine the rate constant and activation energy of the DNA hybridization process. A few simulated models were proposed to represent the structural, mechanical, and kinetic properties of DNA hybridization. The kinetics of strand displacement reactions has also been studied as a special case of DNA hybridization. The thermodynamic and kinetic characteristics of DNA nanostructures have been exploited to develop rapid and isothermal annealing protocols. It is conceivable that a more thorough understanding of the DNA assembly process could be used to guide the structural design process and optimize the conditions for assembly, manipulation, and functionalization, thus benefiting both upstream design and downstream applications.

Guiding the folding pathway of DNA origami

NASA Astrophysics Data System (ADS)

Dunn, Katherine E.; Dannenberg, Frits; Ouldridge, Thomas E.; Kwiatkowska, Marta; Turberfield, Andrew J.; Bath, Jonathan

2015-09-01

DNA origami is a robust assembly technique that folds a single-stranded DNA template into a target structure by annealing it with hundreds of short `staple' strands. Its guiding design principle is that the target structure is the single most stable configuration. The folding transition is cooperative and, as in the case of proteins, is governed by information encoded in the polymer sequence. A typical origami folds primarily into the desired shape, but misfolded structures can kinetically trap the system and reduce the yield. Although adjusting assembly conditions or following empirical design rules can improve yield, well-folded origami often need to be separated from misfolded structures. The problem could in principle be avoided if assembly pathway and kinetics were fully understood and then rationally optimized. To this end, here we present a DNA origami system with the unusual property of being able to form a small set of distinguishable and well-folded shapes that represent discrete and approximately degenerate energy minima in a vast folding landscape, thus allowing us to probe the assembly process. The obtained high yield of well-folded origami structures confirms the existence of efficient folding pathways, while the shape distribution provides information about individual trajectories through the folding landscape. We find that, similarly to protein folding, the assembly of DNA origami is highly cooperative; that reversible bond formation is important in recovering from transient misfoldings; and that the early formation of long-range connections can very effectively enforce particular folds. We use these insights to inform the design of the system so as to steer assembly towards desired structures. Expanding the rational design process to include the assembly pathway should thus enable more reproducible synthesis, particularly when targeting more complex structures. We anticipate that this expansion will be essential if DNA origami is to continue its rapid development and become a reliable manufacturing technology.

Guiding the folding pathway of DNA origami.

PubMed

Dunn, Katherine E; Dannenberg, Frits; Ouldridge, Thomas E; Kwiatkowska, Marta; Turberfield, Andrew J; Bath, Jonathan

2015-09-03

DNA origami is a robust assembly technique that folds a single-stranded DNA template into a target structure by annealing it with hundreds of short 'staple' strands. Its guiding design principle is that the target structure is the single most stable configuration. The folding transition is cooperative and, as in the case of proteins, is governed by information encoded in the polymer sequence. A typical origami folds primarily into the desired shape, but misfolded structures can kinetically trap the system and reduce the yield. Although adjusting assembly conditions or following empirical design rules can improve yield, well-folded origami often need to be separated from misfolded structures. The problem could in principle be avoided if assembly pathway and kinetics were fully understood and then rationally optimized. To this end, here we present a DNA origami system with the unusual property of being able to form a small set of distinguishable and well-folded shapes that represent discrete and approximately degenerate energy minima in a vast folding landscape, thus allowing us to probe the assembly process. The obtained high yield of well-folded origami structures confirms the existence of efficient folding pathways, while the shape distribution provides information about individual trajectories through the folding landscape. We find that, similarly to protein folding, the assembly of DNA origami is highly cooperative; that reversible bond formation is important in recovering from transient misfoldings; and that the early formation of long-range connections can very effectively enforce particular folds. We use these insights to inform the design of the system so as to steer assembly towards desired structures. Expanding the rational design process to include the assembly pathway should thus enable more reproducible synthesis, particularly when targeting more complex structures. We anticipate that this expansion will be essential if DNA origami is to continue its rapid development and become a reliable manufacturing technology.

Transition between Two Regimes Describing Internal Fluctuation of DNA in a Nanochannel

PubMed Central

Su, Tianxiang; Das, Somes K.; Xiao, Ming; Purohit, Prashant K.

2011-01-01

We measure the thermal fluctuation of the internal segments of a piece of DNA confined in a nanochannel about 50100 nm wide. This local thermodynamic property is key to accurate measurement of distances in genomic analysis. For DNA in 100 nm channels, we observe a critical length scale 10 m for the mean extension of internal segments, below which the de Gennes' theory describes the fluctuations with no fitting parameters, and above which the fluctuation data falls into Odijk's deflection theory regime. By analyzing the probability distributions of the extensions of the internal segments, we infer that folded structures of length 150250 nm, separated by 10 m exist in the confined DNA during the transition between the two regimes. For 50 nm channels we find that the fluctuation is significantly reduced since the Odijk regime appears earlier. This is critical for genomic analysis. We further propose a more detailed theory based on small fluctuations and incorporating the effects of confinement to explicitly calculate the statistical properties of the internal fluctuations. Our theory is applicable to polymers with heterogeneous mechanical properties confined in non-uniform channels. We show that existing theories for the end-to-end extension/fluctuation of polymers can be used to study the internal fluctuations only when the contour length of the polymer is many times larger than its persistence length. Finally, our results suggest that introducing nicks in the DNA will not change its fluctuation behavior when the nick density is below 1 nick per kbp DNA. PMID:21423606

Yeast mitochondrial HMG proteins: DNA-binding properties of the most evolutionarily divergent component of mitochondrial nucleoids.

PubMed

Bakkaiova, Jana; Marini, Victoria; Willcox, Smaranda; Nosek, Jozef; Griffith, Jack D; Krejci, Lumir; Tomaska, Lubomir

2015-12-08

Yeast mtDNA is compacted into nucleoprotein structures called mitochondrial nucleoids (mt-nucleoids). The principal mediators of nucleoid formation are mitochondrial high-mobility group (HMG)-box containing (mtHMG) proteins. Although these proteins are some of the fastest evolving components of mt-nucleoids, it is not known whether the divergence of mtHMG proteins on the level of their amino acid sequences is accompanied by diversification of their biochemical properties. In the present study we performed a comparative biochemical analysis of yeast mtHMG proteins from Saccharomyces cerevisiae (ScAbf2p), Yarrowia lipolytica (YlMhb1p) and Candida parapsilosis (CpGcf1p). We found that all three proteins exhibit relatively weak binding to intact dsDNA. In fact, ScAbf2p and YlMhb1p bind quantitatively to this substrate only at very high protein to DNA ratios and CpGcf1p shows only negligible binding to dsDNA. In contrast, the proteins exhibit much higher preference for recombination intermediates such as Holliday junctions (HJ) and replication forks (RF). Therefore, we hypothesize that the roles of the yeast mtHMG proteins in maintenance and compaction of mtDNA in vivo are in large part mediated by their binding to recombination/replication intermediates. We also speculate that the distinct biochemical properties of CpGcf1p may represent one of the prerequisites for frequent evolutionary tinkering with the form of the mitochondrial genome in the CTG-clade of hemiascomycetous yeast species. © 2016 Authors.

RNA polymerase gate loop guides the nontemplate DNA strand in transcription complexes.

PubMed

NandyMazumdar, Monali; Nedialkov, Yuri; Svetlov, Dmitri; Sevostyanova, Anastasia; Belogurov, Georgiy A; Artsimovitch, Irina

2016-12-27

Upon RNA polymerase (RNAP) binding to a promoter, the σ factor initiates DNA strand separation and captures the melted nontemplate DNA, whereas the core enzyme establishes interactions with the duplex DNA in front of the active site that stabilize initiation complexes and persist throughout elongation. Among many core RNAP elements that participate in these interactions, the β' clamp domain plays the most prominent role. In this work, we investigate the role of the β gate loop, a conserved and essential structural element that lies across the DNA channel from the clamp, in transcription regulation. The gate loop was proposed to control DNA loading during initiation and to interact with NusG-like proteins to lock RNAP in a closed, processive state during elongation. We show that the removal of the gate loop has large effects on promoter complexes, trapping an unstable intermediate in which the RNAP contacts with the nontemplate strand discriminator region and the downstream duplex DNA are not yet fully established. We find that although RNAP lacking the gate loop displays moderate defects in pausing, transcript cleavage, and termination, it is fully responsive to the transcription elongation factor NusG. Together with the structural data, our results support a model in which the gate loop, acting in concert with initiation or elongation factors, guides the nontemplate DNA in transcription complexes, thereby modulating their regulatory properties.

Regular square planer bis-(4,4,4-trifluoro-1-(thiophen-2-yl)butane-1,3-dione)/copper(II) complex: Trans/cis-DFT isomerization, crystal structure, thermal, solvatochromism, hirshfeld surface and DNA-binding analysis

NASA Astrophysics Data System (ADS)

Hema, M. K.; Karthik, C. S.; Warad, Ismail; Lokanath, N. K.; Zarrouk, Abdelkader; Kumara, Karthik; Pampa, K. J.; Mallu, P.

2018-04-01

Trans-[Cu(O∩O)2] complex, O∩O = 4,4,4-trifluoro-1-(thiophen-2-yl)butane-1,3-dione was reported with high potential toward CT-DNA binder. The solved XRD-structure of complex indicated a perfect regular square-planer geometry around the Cu(II) center. The trans/cis-DFT-isomerization calculation supported the XRD seen in reflecting the trans-isomer as the kinetic-favor isomer. The desired complex structure was also characterized by conductivity measurement, CHN-elemental analyses, MS, EDX, SEM, UV-Vis., FT-IR, HAS and TG/DTG. The Solvatochromism behavior of the complex was evaluated using four different polar solvents. MPE and Hirshfeld surface analysis (HSA) come to an agreement that fluoride and thiophene protons atoms are with suitable electro-potential environment to form non-classical H-bonds of type CThsbnd H⋯F. The DNA-binding properties were investigated by viscosity tests and spectrometric titrations, the results revealed the complex as strong calf-thymus DNA binder. High intrinsic-binding constants value ∼1.8 × 105 was collected.

Amplified biosensing using the horseradish peroxidase-mimicking DNAzyme as an electrocatalyst.

PubMed

Pelossof, Gilad; Tel-Vered, Ran; Elbaz, Johann; Willner, Itamar

2010-06-01

The hemin/G-quadruplex horseradish peroxidase-mimicking DNAzyme is assembled on Au electrodes. It reveals bioelectrocatalytic properties and electrocatalyzes the reduction of H(2)O(2). The bioelectrocatalytic functions of the hemin/G-quadruplex DNAzyme are used to develop electrochemical sensors that follow the activity of glucose oxidase and biosensors for the detection of DNA or low-molecular-weight substrates (adenosine monophosphate, AMP). Hairpin nucleic structures that include the G-quadruplex sequence in a caged configuration and the nucleic acid sequence complementary to the analyte DNA, or the aptamer sequence for AMP, are immobilized on Au-electrode surfaces. In the presence of the DNA analyte, or AMP, the hairpin structures are opened, and the hemin/G-quadruplex horseradish peroxidase-mimicking DNAzyme structures are generated on the electrode surfaces. The bioelectrocatalytic cathodic currents generated by the functionalized electrodes, upon the electrochemical reduction of H(2)O(2), provide a quantitative measure for the detection of the target analytes. The DNA target was analyzed with a detection limit of 1 x 10(-12) M, while the detection limit for analyzing AMP was 1 x 10(-6) M. Methods to regenerate the sensing surfaces are presented.

DNA-Mediated Self-Organization of Polymeric Nanocompartments Leads to Interconnected Artificial Organelles.

PubMed

Liu, Juan; Postupalenko, Viktoriia; Lörcher, Samuel; Wu, Dalin; Chami, Mohamed; Meier, Wolfgang; Palivan, Cornelia G

2016-11-09

Self-organization of nanocomponents was mainly focused on solid nanoparticles, quantum dots, or liposomes to generate complex architectures with specific properties, but intrinsically limited or not developed enough, to mimic sophisticated structures with biological functions in cells. Here, we present a biomimetic strategy to self-organize synthetic nanocompartments (polymersomes) into clusters with controlled properties and topology by exploiting DNA hybridization to interconnect polymersomes. Molecular and external factors affecting the self-organization served to design clusters mimicking the connection of natural organelles: fine-tune of the distance between tethered polymersomes, different topologies, no fusion of clustered polymersomes, and no aggregation. Unexpected, extended DNA bridges that result from migration of the DNA strands inside the thick polymer membrane (about 12 nm) represent a key stability and control factor, not yet exploited for other synthetic nano-object networks. The replacement of the empty polymersomes with artificial organelles, already reported for single polymersome architecture, will provide an excellent platform for the development of artificial systems mimicking natural organelles or cells and represents a fundamental step in the engineering of molecular factories.

Sequence requirements of oligonucleotide chiral selectors for the capillary electrophoresis resolution of low-affinity DNA binders.

PubMed

Tohala, Luma; Oukacine, Farid; Ravelet, Corinne; Peyrin, Eric

2017-05-01

We recently reported that a great variety of DNA oligonucleotides (ONs) used as chiral selectors in partial-filling capillary electrophoresis (CE) exhibited interesting enantioresolution properties toward low-affinity DNA binders. Herein, the sequence prerequisites of ONs for the CE enantioseparation process were studied. First, the chiral resolution properties of a series of homopolymeric sequences (Poly-dT) of different lengths (from 5 to 60-mer) were investigated. It was shown that the size increase-dependent random coil-like conformation of Poly-dT favorably acted on the apparent selectivity and resolution. The base-unpairing state constituted also an important factor in the chiral resolution ability of ONs as the switch from the single-stranded to double-stranded structure was responsible for a significant decrease in the analyte selectivity range. Finally, the chemical diversity enhanced the enantioresolution ability of single-stranded ONs. The present work could lay the foundation for the design of performant ON chiral selectors for the CE separation of weak DNA binder enantiomers. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

People’s Republic of China Scientific Abstracts, Number 197

DTIC Science & Technology

1978-08-10

Chinese Academy of Sciences TITLE: "Characteristics of the DNA from Pupal Gonads of Bombyx Mori L.1*» SOURCE: Peking K’UN-CH’UNG HSUEH-PAO [ACTA...some fundamental properties of the DNA from the pupal gonada of Bombyx mori L. The results are as follows: The peak of u.v. absorption of the testicular...pest populations, especially by means of using their natural enemies and changing the spatial structures of their habitats ; (4) use of descrip

Recognition of the DNA sequence by an inorganic crystal surface

PubMed Central

Sampaolese, Beatrice; Bergia, Anna; Scipioni, Anita; Zuccheri, Giampaolo; Savino, Maria; Samorì, Bruno; De Santis, Pasquale

2002-01-01

The sequence-dependent curvature is generally recognized as an important and biologically relevant property of DNA because it is involved in the formation and stability of association complexes with proteins. When a DNA tract, intrinsically curved for the periodical recurrence on the same strand of A-tracts phased with the B-DNA periodicity, is deposited on a flat surface, it exposes to that surface either a T- or an A-rich face. The surface of a freshly cleaved mica crystal recognizes those two faces and preferentially interacts with the former one. Statistical analysis of scanning force microscopy (SFM) images provides evidence of this recognition between an inorganic crystal surface and nanoscale structures of double-stranded DNA. This finding could open the way toward the use of the sequence-dependent adhesion to specific crystal faces for nanotechnological purposes. PMID:12361979

Light-driven enzymatic catalysis of DNA repair: a review of recent biophysical studies on photolyase.

PubMed

Weber, Stefan

2005-02-25

More than 50 years ago, initial experiments on enzymatic photorepair of ultraviolet (UV)-damaged DNA were reported [Proc. Natl. Acad. Sci. U. S. A. 35 (1949) 73]. Soon after this discovery, it was recognized that one enzyme, photolyase, is able to repair UV-induced DNA lesions by effectively reversing their formation using blue light. The enzymatic process named DNA photoreactivation depends on a non-covalently bound cofactor, flavin adenine dinucleotide (FAD). Flavins are ubiquitous redox-active catalysts in one- and two-electron transfer reactions of numerous biological processes. However, in the case of photolyase, not only the ground-state redox properties of the FAD cofactor are exploited but also, and perhaps more importantly, its excited-state properties. In the catalytically active, fully reduced redox form, the FAD absorbs in the blue and near-UV ranges of visible light. Although there is no direct experimental evidence, it appears generally accepted that starting from the excited singlet state, the chromophore initiates a reductive cleavage of the two major DNA photodamages, cyclobutane pyrimidine dimers and (6-4) photoproducts, by short-distance electron transfer to the DNA lesion. Back electron transfer from the repaired DNA segment is believed to eventually restore the initial redox states of the cofactor and the DNA nucleobases, resulting in an overall reaction with net-zero exchanged electrons. Thus, the entire process represents a true catalytic cycle. Many biochemical and biophysical studies have been carried out to unravel the fundamentals of this unique mode of action. The work has culminated in the elucidation of the three-dimensional structure of the enzyme in 1995 that revealed remarkable details, such as the FAD-cofactor arrangement in an unusual U-shaped configuration. With the crystal structure of the enzyme at hand, research on photolyases did not come to an end but, for good reason, intensified: the geometrical structure of the enzyme alone is not sufficient to fully understand the enzyme's action on UV-damaged DNA. Much effort has therefore been invested to learn more about, for example, the geometry of the enzyme-substrate complex, and the mechanism and pathways of intra-enzyme and enzyme <-->DNA electron transfer. Many of the key results from biochemical and molecular biology characterizations of the enzyme or the enzyme-substrate complex have been summarized in a number of reviews. Complementary to these articles, this review focuses on recent biophysical studies of photoreactivation comprising work performed from the early 1990s until the present.

NMR structure of the DNA decamer duplex containing double T*G mismatches of cis-syn cyclobutane pyrimidine dimer: implications for DNA damage recognition by the XPC-hHR23B complex.

PubMed

Lee, Joon-Hwa; Park, Chin-Ju; Shin, Jae-Sun; Ikegami, Takahisa; Akutsu, Hideo; Choi, Byong-Seok

2004-01-01

The cis-syn cyclobutane pyrimidine dimer (CPD) is a cytotoxic, mutagenic and carcinogenic DNA photoproduct and is repaired by the nucleotide excision repair (NER) pathway in mammalian cells. The XPC-hHR23B complex as the initiator of global genomic NER binds to sites of certain kinds of DNA damage. Although CPDs are rarely recognized by the XPC-hHR23B complex, the presence of mismatched bases opposite a CPD significantly increased the binding affinity of the XPC-hHR23B complex to the CPD. In order to decipher the properties of the DNA structures that determine the binding affinity for XPC-hHR23B to DNA, we carried out structural analyses of the various types of CPDs by NMR spectroscopy. The DNA duplex which contains a single 3' T*G wobble pair in a CPD (CPD/GA duplex) induces little conformational distortion. However, severe distortion of the helical conformation occurs when a CPD contains double T*G wobble pairs (CPD/GG duplex) even though the T residues of the CPD form stable hydrogen bonds with the opposite G residues. The helical bending angle of the CPD/GG duplex was larger than those of the CPD/GA duplex and properly matched CPD/AA duplex. The fluctuation of the backbone conformation and significant changes in the widths of the major and minor grooves at the double T*G wobble paired site were also observed in the CPD/GG duplex. These structural features were also found in a duplex that contains the (6-4) adduct, which is efficiently recognized by the XPC-hHR23B complex. Thus, we suggest that the unique structural features of the DNA double helix (that is, helical bending, flexible backbone conformation, and significant changes of the major and/or minor grooves) might be important factors in determining the binding affinity of the XPC-hHR23B complex to DNA.

Casting inorganic structures with DNA molds

PubMed Central

Sun, Wei; Boulais, Etienne; Hakobyan, Yera; Wang, Wei Li; Guan, Amy; Bathe, Mark; Yin, Peng

2014-01-01

We report a general strategy for designing and synthesizing inorganic nanostructures with arbitrarily prescribed three-dimensional shapes. Computationally designed DNA strands self-assemble into a stiff “nano-mold” that contains a user-specified three-dimensional cavity and encloses a nucleating gold “seed”. Under mild conditions, this seed grows into a larger cast structure that fills and thus replicates the cavity. We synthesized a variety of nanoparticles with three nanometer resolution: three distinct silver cuboids with three independently tunable dimensions, silver and gold nanoparticles with diverse cross sections, and composite structures with homo-/heterogeneous components. The designer equilateral silver triangular and spherical nanoparticles exhibited plasmonic properties consistent with electromagnetism-based simulations. Our framework is generalizable to more complex geometries and diverse inorganic materials, offering a range of applications in biosensing, photonics, and nanoelectronics. PMID:25301973

Modeling of DNA local parameters predicts encrypted architectural motifs in Xenopus laevis ribosomal gene promoter

PubMed Central

Roux-Rouquie, Magali; Marilley, Monique

2000-01-01

We have modeled local DNA sequence parameters to search for DNA architectural motifs involved in transcription regulation and promotion within the Xenopus laevis ribosomal gene promoter and the intergenic spacer (IGS) sequences. The IGS was found to be shaped into distinct topological domains. First, intrinsic bends split the IGS into domains of common but different helical features. Local parameters at inter-domain junctions exhibit a high variability with respect to intrinsic curvature, bendability and thermal stability. Secondly, the repeated sequence blocks of the IGS exhibit right-handed supercoiled structures which could be related to their enhancer properties. Thirdly, the gene promoter presents both inherent curvature and minor groove narrowing which may be viewed as motifs of a structural code for protein recognition and binding. Such pre-existing deformations could simply be remodeled during the binding of the transcription complex. Alternatively, these deformations could pre-shape the promoter in such a way that further remodeling is facilitated. Mutations shown to abolish promoter curvature as well as intrinsic minor groove narrowing, in a variant which maintained full transcriptional activity, bring circumstantial evidence for structurally-preorganized motifs in relation to transcription regulation and promotion. Using well documented X.laevis rDNA regulatory sequences we showed that computer modeling may be of invaluable assistance in assessing encrypted architectural motifs. The evidence of these DNA topological motifs with respect to the concept of structural code is discussed. PMID:10982860

Interactions of Ku70/80 with Double-Strand DNA: Energetic, Dynamics, and Functional Implications

NASA Technical Reports Server (NTRS)

Hu, Shaowen; Cucinotta, Francis A.

2010-01-01

Space radiation is a proficient inducer of DNA damage leading to mutation, aberrant cell signaling, and cancer formation. Ku is among the first responding proteins in nucleus to recognize and bind the DNA double strand breaks (DSBs) whenever they are introduced. Once loaded Ku works as a scaffold to recruit other repair factors of non-homologous end joining and facilitates the following repair processes. The crystallographic study of the Ku70/80 heterodimer indicate the core structure of this protein shows virtually no conformational change after binding with DNA. To investigate the dynamical features as well as the energetic characteristics of Ku-DNA binding, we conduct multi-nanosecond molecular dynamics simulations of a modeled Ku70/80 structure and several complexes with two 24-bp DNA duplexes. Free energy calculations show significant energy differences between the complexes with Ku bound at DSBs and those with Ku associated at an internal site of a chromosome. The results also reveal detailed interactions between different nucleotides and the amino acids along the DNA-binding cradle of Ku, indicating subtle binding preference of Ku at specific DNA sequences. The covariance matrix analyses along the trajectories demonstrate the protein is stimulated to undergo correlated motions of different domains once bound to DNA ends. Additionally, principle component analyses identify these low frequency collective motions suitable for binding with and translocation along duplex DNA. It is proposed that the modification of dynamical properties of Ku upon binding with DSBs may provide a signal for the further recruitment of other repair factors such as DNA-PKcs, XLF, and XRCC4.

Study of intermolecular contacts in the proline-rich homeodomain (PRH)-DNA complex using molecular dynamics simulations.

PubMed

Jalili, Seifollah; Karami, Leila

2012-03-01

The proline-rich homeodomain (PRH)-DNA complex consists of a protein with 60 residues and a 13-base-pair DNA. The PRH protein is a transcription factor that plays a key role in the regulation of gene expression. PRH is a significant member of the Q50 class of homeodomain proteins. The homeodomain section of PRH is essential for binding to DNA and mediates sequence-specific DNA binding. Three 20-ns molecular dynamics (MD) simulations (free protein, free DNA and protein-DNA complex) in explicit solvent water were performed to elucidate the intermolecular contacts in the PRH-DNA complex and the role of dynamics of water molecules forming water-mediated contacts. The simulation provides a detailed explanation of the trajectory of hydration water molecules. The simulations show that some water molecules in the protein-DNA interface exchange with bulk waters. The simulation identifies that most of the contacts consisted of direct interactions between the protein and DNA including specific and non-specific contacts, but several water-mediated polar contacts were also observed. The specific interaction between Gln50 and C18 and water-mediated hydrogen bond between Gln50 and T7 were found to be present during almost the entire time of the simulation. These results show good consistency with experimental and previous computational studies. Structural properties such as root-mean-square deviations (RMSD), root-mean-square fluctuations (RMSF) and secondary structure were also analyzed as a function of time. Analyses of the trajectories showed that the dynamic fluctuations of both the protein and the DNA were lowered by the complex formation.

BIOPHYSICAL PROPERTIES OF NUCLEIC ACIDS AT SURFACES RELEVANT TO MICROARRAY PERFORMANCE

PubMed Central

Rao, Archana N.; Grainger, David W.

2014-01-01

Both clinical and analytical metrics produced by microarray-based assay technology have recognized problems in reproducibility, reliability and analytical sensitivity. These issues are often attributed to poor understanding and control of nucleic acid behaviors and properties at solid-liquid interfaces. Nucleic acid hybridization, central to DNA and RNA microarray formats, depends on the properties and behaviors of single strand (ss) nucleic acids (e.g., probe oligomeric DNA) bound to surfaces. ssDNA’s persistence length, radius of gyration, electrostatics, conformations on different surfaces and under various assay conditions, its chain flexibility and curvature, charging effects in ionic solutions, and fluorescent labeling all influence its physical chemistry and hybridization under assay conditions. Nucleic acid (e.g., both RNA and DNA) target interactions with immobilized ssDNA strands are highly impacted by these biophysical states. Furthermore, the kinetics, thermodynamics, and enthalpic and entropic contributions to DNA hybridization reflect global probe/target structures and interaction dynamics. Here we review several biophysical issues relevant to oligomeric nucleic acid molecular behaviors at surfaces and their influences on duplex formation that influence microarray assay performance. Correlation of biophysical aspects of single and double-stranded nucleic acids with their complexes in bulk solution is common. Such analysis at surfaces is not commonly reported, despite its importance to microarray assays. We seek to provide further insight into nucleic acid-surface challenges facing microarray diagnostic formats that have hindered their clinical adoption and compromise their research quality and value as genomics tools. PMID:24765522

Engineering nucleic acid structures for programmable molecular circuitry and intracellular biocomputation

NASA Astrophysics Data System (ADS)

Li, Jiang; Green, Alexander A.; Yan, Hao; Fan, Chunhai

2017-11-01

Nucleic acids have attracted widespread attention due to the simplicity with which they can be designed to form discrete structures and programmed to perform specific functions at the nanoscale. The advantages of DNA/RNA nanotechnology offer numerous opportunities for in-cell and in-vivo applications, and the technology holds great promise to advance the growing field of synthetic biology. Many elegant examples have revealed the potential in integrating nucleic acid nanostructures in cells and in vivo where they can perform important physiological functions. In this Review, we summarize the current abilities of DNA/RNA nanotechnology to realize applications in live cells and then discuss the key problems that must be solved to fully exploit the useful properties of nanostructures. Finally, we provide viewpoints on how to integrate the tools provided by DNA/RNA nanotechnology and related new technologies to construct nucleic acid nanostructure-based molecular circuitry for synthetic biology.

Effect of Methylation on Local Mechanics and Hydration Structure of DNA.

PubMed

Teng, Xiaojing; Hwang, Wonmuk

2018-04-24

Cytosine methylation affects mechanical properties of DNA and potentially alters the hydration fingerprint for recognition by proteins. The atomistic origin for these effects is not well understood, and we address this via all-atom molecular dynamics simulations. We find that the stiffness of the methylated dinucleotide step changes marginally, whereas the neighboring steps become stiffer. Stiffening is further enhanced for consecutively methylated steps, providing a mechanistic origin for the effect of hypermethylation. Steric interactions between the added methyl groups and the nonpolar groups of the neighboring nucleotides are responsible for the stiffening in most cases. By constructing hydration maps, we found that methylation also alters the surface hydration structure in distinct ways. Its resistance to deformation may contribute to the stiffening of DNA for deformational modes lacking steric interactions. These results highlight the sequence- and deformational-mode-dependent effects of cytosine methylation. Copyright © 2018 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Interplay of protein and DNA structure revealed in simulations of the lac operon.

PubMed

Czapla, Luke; Grosner, Michael A; Swigon, David; Olson, Wilma K

2013-01-01

The E. coli Lac repressor is the classic textbook example of a protein that attaches to widely spaced sites along a genome and forces the intervening DNA into a loop. The short loops implicated in the regulation of the lac operon suggest the involvement of factors other than DNA and repressor in gene control. The molecular simulations presented here examine two likely structural contributions to the in-vivo looping of bacterial DNA: the distortions of the double helix introduced upon association of the highly abundant, nonspecific nucleoid protein HU and the large-scale deformations of the repressor detected in low-resolution experiments. The computations take account of the three-dimensional arrangements of nucleotides and amino acids found in crystal structures of DNA with the two proteins, the natural rest state and deformational properties of protein-free DNA, and the constraints on looping imposed by the conformation of the repressor and the orientation of bound DNA. The predicted looping propensities capture the complex, chain-length-dependent variation in repression efficacy extracted from gene expression studies and in vitro experiments and reveal unexpected chain-length-dependent variations in the uptake of HU, the deformation of repressor, and the folding of DNA. Both the opening of repressor and the presence of HU, at levels approximating those found in vivo, enhance the probability of loop formation. HU affects the global organization of the repressor and the opening of repressor influences the levels of HU binding to DNA. The length of the loop determines whether the DNA adopts antiparallel or parallel orientations on the repressor, whether the repressor is opened or closed, and how many HU molecules bind to the loop. The collective behavior of proteins and DNA is greater than the sum of the parts and hints of ways in which multiple proteins may coordinate the packaging and processing of genetic information.

Nickel-quinolones interaction. Part 4. Structure and biological evaluation of nickel(II)-enrofloxacin complexes compared to zinc(II) analogues.

PubMed

Skyrianou, Kalliopi C; Psycharis, Vassilis; Raptopoulou, Catherine P; Kessissoglou, Dimitris P; Psomas, George

2011-01-01

The nickel(II) complexes with the second-generation quinolone antibacterial agent enrofloxacin in the presence or absence of the nitrogen-donor heterocyclic ligands 1,10-phenanthroline, 2,2'-bipyridine or pyridine have been synthesized and characterized. Enrofloxacin acts as bidentate ligand coordinated to Ni(II) ion through the ketone oxygen and a carboxylato oxygen. The crystal structure of (1,10-phenanthroline)bis(enrofloxacinato)nickel(II) has been determined by X-ray crystallography. UV study of the interaction of the complexes with calf-thymus DNA (CT DNA) has shown that they bind to CT DNA and bis(pyridine)bis(enrofloxacinato)nickel(II) exhibits the highest binding constant to CT DNA. The cyclic voltammograms of the complexes have shown that in the presence of CT DNA the complexes can bind to CT DNA by the intercalative binding mode which has also been verified by DNA solution viscosity measurements. Competitive study with ethidium bromide (EB) has shown that the complexes can displace the DNA-bound EB indicating that they bind to DNA in strong competition with EB. The complexes exhibit good binding propensity to human or bovine serum albumin protein having relatively high binding constant values. The biological properties of the complexes have been evaluated in comparison to the corresponding Zn(II) enrofloxacinato complexes as well as Ni(II) complexes with the first-generation quinolone oxolinic acid. Copyright © 2010 Elsevier Inc. All rights reserved.

Release of the cyano moiety in the crystal structure of N-cyanomethyl-N-(2-methoxyethyl)-daunomycin complexed with d(CGATCG).

PubMed

Saminadin, P; Dautant, A; Mondon, M; Langlois D'estaintot, B; Courseille, C; Précigoux, G

2000-01-01

Doxorubicin is among the most widely used anthracycline in cancer chemotherapy. In an attempt to avoid the cardiotoxicity and drug resistance of doxorubicin therapy, several analogues were synthesized. The cyanomorpholinyl derivative is the most cytotoxic. They differ greatly from their parent compound in their biological and pharmacological properties, inducing cross-links in drug DNA complexes. The present study concerns N-cyanomethyl-N-(2-methoxyethyl)-daunomycin (CMDa), a synthetic analogue of cyanomorpholino-daunomycin. Compared to doxorubicin, CMDa displays a cytotoxic activity on L1210 leukemia cells at higher concentration but is effective on doxorubicin resistant cells. The results of fluorescence quenching experiments as well as the melting temperature (DeltaTm = 7.5 degrees C) studies are consistent with a drug molecule which intercalates between the DNA base pairs and stabilizes the DNA double helix. The crystal structure of CMDa complexed to the hexanucleotide d(CGATCG) has been determined at 1.5 A resolution. The complex crystallizes in the space group P41212 and is similar to other anthracycline-hexanucleotide complexes. In the crystal state, the observed densities indicate the formation of N-hydroxymethyl-N-(2-methoxyethyl)-daunomycin (HMDa) with the release of the cyano moiety without DNA alkylation. The formation of this degradation compound is discussed in relation with other drug modifications when binding to DNA. Comparison with two other drug-DNA crystal structures suggests a correlation between a slight change in DNA conformation and the nature of the amino sugar substituents at the N3' position located in the minor groove.

Hybrid ligand-alkylating agents targeting telomeric G-quadruplex structures.

PubMed

Doria, Filippo; Nadai, Matteo; Folini, Marco; Di Antonio, Marco; Germani, Luca; Percivalle, Claudia; Sissi, Claudia; Zaffaroni, Nadia; Alcaro, Stefano; Artese, Anna; Richter, Sara N; Freccero, Mauro

2012-04-14

The synthesis, physico-chemical properties and biological effects of a new class of naphthalene diimides (NDIs) capable of reversibly binding telomeric DNA and alkylate it through an electrophilic quinone methide moiety (QM), are reported. FRET and circular dichroism assays showed a marked stabilization and selectivity towards telomeric G4 DNA folded in a hybrid topology. NDI-QMs' alkylating properties revealed a good reactivity on single nucleosides and selectivity towards telomeric G4. A selected NDI was able to significantly impair the growth of melanoma cells by causing telomere dysfunction and down-regulation of telomerase expression. These findings points to our hybrid ligand-alkylating NDIs as possible tools for the development of novel targeted anticancer therapies. This journal is © The Royal Society of Chemistry 2012

Evaluation of methods for the extraction of DNA from drinking water distribution system biofilms.

PubMed

Hwang, Chiachi; Ling, Fangqiong; Andersen, Gary L; LeChevallier, Mark W; Liu, Wen-Tso

2012-01-01

While drinking water biofilms have been characterized in various drinking water distribution systems (DWDS), little is known about the impact of different DNA extraction methods on the subsequent analysis of microbial communities in drinking water biofilms. Since different DNA extraction methods have been shown to affect the outcome of microbial community analysis in other environments, it is necessary to select a DNA extraction method prior to the application of molecular tools to characterize the complex microbial ecology of the DWDS. This study compared the quantity and quality of DNA yields from selected DWDS bacteria with different cell wall properties using five widely used DNA extraction methods. These were further selected and evaluated for their efficiency and reproducibility of DNA extraction from DWDS samples. Terminal restriction fragment length analysis and the 454 pyrosequencing technique were used to interpret the differences in microbial community structure and composition, respectively, from extracted DNA. Such assessments serve as a concrete step towards the determination of an optimal DNA extraction method for drinking water biofilms, which can then provide a reliable comparison of the meta-analysis results obtained in different laboratories.

A nucleotide binding rectification Brownian ratchet model for translocation of Y-family DNA polymerases

PubMed Central

2011-01-01

Y-family DNA polymerases are characterized by low-fidelity synthesis on undamaged DNA and ability to catalyze translesion synthesis over the damaged DNA. Their translocation along the DNA template is an important event during processive DNA synthesis. In this work we present a Brownian ratchet model for this translocation, where the directed translocation is rectified by the nucleotide binding to the polymerase. Using the model, different features of the available structures for Dpo4, Dbh and polymerase ι in binary and ternary forms can be easily explained. Other dynamic properties of the Y-family polymerases such as the fast translocation event upon dNTP binding for Dpo4 and the considerable variations of the processivity among the polymerases can also be well explained by using the model. In addition, some predicted results of the DNA synthesis rate versus the external force acting on Dpo4 and Dbh polymerases are presented. Moreover, we compare the effect of the external force on the DNA synthesis rate of the Y-family polymerase with that of the replicative DNA polymerase. PMID:21699732

The meta-epigenomic structure of purified human stem cell populations is defined at cis-regulatory sequences

PubMed Central

Zhao, Yong Mei; Golden, Aaron; Mar, Jessica C.; Einstein, Francine H.; Greally, John M.

2014-01-01

The mechanism and significance of epigenetic variability in the same cell type between healthy individuals are not clear. Here, we purify human CD34+ hematopoietic stem and progenitor cells (HSPCs) from different individuals and find that there is increased variability of DNA methylation at loci with properties of promoters and enhancers. The variability is especially enriched at candidate enhancers near genes transitioning between silent and expressed states, and encoding proteins with leukocyte differentiation properties. Our findings of increased variability at loci with intermediate DNA methylation values, at candidate “poised” enhancers, and at genes involved in HSPC lineage commitment suggest that CD34+ cell subtype heterogeneity between individuals is a major mechanism for the variability observed. Epigenomic studies performed on cell populations, even when purified, are testing collections of epigenomes, or meta-epigenomes. Our findings show that meta-epigenomic approaches to data analysis can provide insights into cell subpopulation structure. PMID:25327398

Structural and biomechanical characterizations of porcine myocardial extracellular matrix

PubMed Central

Wang, Bo; Tedder, Mary E.; Perez, Clara E.; Wang, Guangjun; de Jongh Curry, Amy L.; To, Filip; Elder, Steven H.; Williams, Lakiesha N.; Simionescu, Dan T.; Liao, Jun

2012-01-01

Extracellular matrix (ECM) of myocardium plays an important role to maintain a multilayered helical architecture of cardiomyocytes. In this study, we have characterized the structural and biomechanical properties of porcine myocardial ECM. Fresh myocardium were decellularized in a rotating bioreactor using 0.1 % sodium dodecyl sulfate solution. Masson’s trichrome staining and SEM demonstrated the removal of cells and preservation of the interconnected 3D cardiomyocyte lacunae. Movat’s pentachrome staining showed the preservation of cardiac elastin ultrastructure and vascular elastin distribution/alignment. DNA assay result confirmed a 98.59 % reduction in DNA content; the acellular myocardial scaffolds were found completely lack of staining for the porcine α-Gal antigen; and the accelerating enzymatic degradation assessment showed a constant degradation rate. Tensile and shear properties of the acellular myocardial scaffolds were also evaluated. Our observations showed that the acellular myocardial ECM possessed important traits of biodegradable scaffolds, indicating the potentials in cardiac regeneration and whole heart tissue engineering. PMID:22584822

Structural and functional properties of the HIV-1 RNA-tRNA(Lys)3 primer complex annealed by the nucleocapsid protein: comparison with the heat-annealed complex.

PubMed Central

Brulé, Fabienne; Marquet, Roland; Rong, Liwei; Wainberg, Mark A; Roques, Bernard P; Le Grice, Stuart F J; Ehresmann, Bernard; Ehresmann, Chantal

2002-01-01

The conversion of the single-stranded RNA genome into double-stranded DNA by virus-coded reverse transcriptase (RT) is an essential step of the retrovirus life cycle. In human immunodeficiency virus type 1 (HIV-1), RT uses the cellular tRNA(Lys)3 to initiate the (-) strand DNA synthesis. Placement of the primer tRNA(Lys)3 involves binding of its 3'-terminal 18 nt to a complementary region of genomic RNA termed PBS. However, the PBS sequence is not the unique determinant of primer usage and additional contacts are important. This placement is believed to be achieved in vivo by the nucleocapsid domain of Gag or by the mature protein NCp. Up to now, structural information essentially arose from heat-annealed primer-template complexes (Isel et al., J Mol Biol, 1995, 247:236-250; Isel et al., EMBO J, 1999, 18:1038-1048). Here, we investigated the formation of the primer-template complex mediated by NCp and compared structural and functional properties of heat- and NCp-annealed complexes. We showed that both heat- and NCp-mediated procedures allow comparable high yields of annealing. Then, we investigated structural features of both kinds of complexes by enzymatic probing, and we compared their relative efficiency in (-) strong stop DNA synthesis. We did not find any significant differences between these complexes, suggesting that information derived from the heat-annealed complex can be transposed to the NCp-mediated complex and most likely to complexes formed in vivo. PMID:11873759

In silico characterization and analysis of RTBP1 and NgTRF1 protein through MD simulation and molecular docking - A comparative study.

PubMed

Mukherjee, Koel; Pandey, Dev Mani; Vidyarthi, Ambarish Saran

2015-02-06

Gaining access to sequence and structure information of telomere binding proteins helps in understanding the essential biological processes involve in conserved sequence specific interaction between DNA and the proteins. Rice telomere binding protein (RTBP1) and Nicotiana glutinosa telomere repeat binding factor (NgTRF1) are helix turn helix motif type of proteins that plays role in telomeric DNA protection and length regulation. Both the proteins share same type of domain but till now there is very less communication on the in silico studies of these complete proteins.Here we intend to do a comparative study between two proteins through modeling of the complete proteins, physiochemical characterization, MD simulation and DNA-protein docking. I-TASSER and CLC protein work bench was performed to find out the protein 3D structure as well as the different parameters to characterize the proteins. MD simulation was completed by GROMOS forcefield of GROMACS for 10 ns of time stretch. The simulated 3D structures were docked with template DNA (3D DNA modeled through 3D-DART) of TTTAGGG conserved sequence motif using HADDOCK web server.Digging up all the facts about the proteins it was reveled that around 120 amino acids in the tail part was showing a good sequence similarity between the proteins. Molecular modeling, sequence characterization and secondary structure prediction also indicates the similarity between the protein's structure and sequence. The result of MD simulation highlights on the RMSD, RMSF, Rg, PCA and Energy plots which also conveys the similar type of motional behavior between them. The best complex formation for both the proteins in docking result also indicates for the first interaction site which is mainly the helix3 region of the DNA binding domain. The overall computational analysis reveals that RTBP1 and NgTRF1 proteins display good amount of similarity in their physicochemical properties, structure, dynamics and binding mode.

In Silico Characterization and Analysis of RTBP1 and NgTRF1 Protein Through MD Simulation and Molecular Docking: A Comparative Study.

PubMed

Mukherjee, Koel; Pandey, Dev Mani; Vidyarthi, Ambarish Saran

2015-09-01

Gaining access to sequence and structure information of telomere-binding proteins helps in understanding the essential biological processes involve in conserved sequence-specific interaction between DNA and the proteins. Rice telomere-binding protein (RTBP1) and Nicotiana glutinosa telomere repeat binding factor (NgTRF1) are helix-turn-helix motif type of proteins that plays role in telomeric DNA protection and length regulation. Both the proteins share same type of domain, but till now there is very less communication on the in silico studies of these complete proteins. Here we intend to do a comparative study between two proteins through modeling of the complete proteins, physiochemical characterization, MD simulation and DNA-protein docking. I-TASSER and CLC protein work bench was performed to find out the protein 3D structure as well as the different parameters to characterize the proteins. MD simulation was completed by GROMOS forcefield of GROMACS for 10 ns of time stretch. The simulated 3D structures were docked with template DNA (3D DNA modeled through 3D-DART) of TTTAGGG conserved sequence motif using HADDOCK Web server. By digging up all the facts about the proteins, it was revealed that around 120 amino acids in the tail part were showing a good sequence similarity between the proteins. Molecular modeling, sequence characterization and secondary structure prediction also indicate the similarity between the protein's structure and sequence. The result of MD simulation highlights on the RMSD, RMSF, Rg, PCA and energy plots which also conveys the similar type of motional behavior between them. The best complex formation for both the proteins in docking result also indicates for the first interaction site which is mainly the helix3 region of the DNA-binding domain. The overall computational analysis reveals that RTBP1 and NgTRF1 proteins display good amount of similarity in their physicochemical properties, structure, dynamics and binding mode.

Structural polymorphism exhibited by a quasipalindrome present in the locus control region (LCR) of the human beta-globin gene cluster.

PubMed

Kaushik, Mahima; Kukreti, Shrikant

2006-01-01

Structural polymorphism of DNA is a widely accepted property. A simple addition to this perception has been our recent finding, where a single nucleotide polymorphism (SNP) site present in a quasipalindromic sequence of beta-globin LCR exhibited a hairpin-duplex equilibrium. Our current studies explore that secondary structures adopted by individual complementary strands compete with formation of a perfect duplex. Using gel-electrophoresis, ultraviolet (UV)-thermal denaturation, circular dichroism (CD) techniques, we have demonstrated the structural transitions within a perfect duplex containing 11 bp quasipalindromic stretch (TGGGG(G/C)CCCCA), to hairpins and bulge duplex forms. The extended version of the 11 bp duplex, flanked by 5 bp on both sides also demonstrated conformational equilibrium between duplex and hairpin species. Gel-electrophoresis confirms that the duplex coexists with hairpin and bulge duplex/cruciform species. Further, in CD spectra of duplexes, presence of two overlapping positive peaks at 265 and 285 nm suggest the features of A- as well as B-type DNA conformation and show oligomer concentration dependence, manifested in A --> B transition. This indicates the possibility of an architectural switching at quasipalindromic region between linear duplex to a cruciform structure. Such DNA structural variations are likely to be found in the mechanics of molecular recognition and manipulation by proteins.

Metallization of Self-Assembled DNA Templates for Electronic Circuit Fabrication

NASA Astrophysics Data System (ADS)

Uprety, Bibek

This work examines the deposition of metallic and semiconductor elements onto self-assembled DNA templates for the fabrication of nanodevices. Biological molecules like DNA self-assemble into a variety of 2- and 3-D architectures without the need for patterning tools. The templates can also be designed to controllably place functional nanomaterials with molecular precision. These characteristics make DNA an attractive template for fabricating electronic circuits. However, electrically conductive structures are needed for electronic applications. While metallized DNA nanostructures have been demonstrated, the ability to make thin, continuous wires that are electrically conductive still represents a formidable challenge. DNA-templated wires have generally been granular in appearance with a resistivity approximately two to three orders of magnitude higher than that of the bulk material. An improved method for the metallization of DNA origami is examined in this work that addresses these challenges of size, morphology and conductivity of the metallized structure. Specifically, we demonstrated a metallization process that uses gold nanorod seeds followed by anisotropic electroless (autocatalytic) plating to provide improved morphology and greater control of the final metallized width of conducting metal lines. Growth during electroless deposition occurs preferentially in the length direction at a rate that is approximately four times the growth rate in the width direction, which enables fabrication of narrow, continuous wires. The electrical properties of 49 nanowires with widths ranging from 13 nm to 29 nm were characterized, and resistivity values as low as 8.9 x 10-7 -m were measured, which represent some of the smallest nanowires and the lowest resistivity values reported in the literature. The metallization procedure developed on smaller templates was also successfully applied to metallize bigger DNA templates of tens of micrometers in length. In addition, a polymer-assisted annealing process was discovered to possibly improve the resistivity of DNA metal nanowires. Following metallization of bigger DNA origami structures, controlled placement of nanorods on a DNA breadboard to make rectangular, square and T-shaped metallic structures was also demonstrated. For site-specific placement, we modified the surface of the gold nanorods with single-stranded DNA. The rods were then attached to DNA templates via complementary base-pairing between the DNA on the nanorods and the attachment strands engineered into the DNA "breadboard" template. Gaps between the nanorods were then filled controllably via anisotropic plating to make 10 nm diameter continuous metallic structures. Finally, controlled placement of metal (gold) - semiconductor (tellurium) materials on a single DNA origami template was demonstrated. The combination of molecularly directed deposition of different nanomaterials and anisotropic metallization presented in this work represents important progress towards the creation of nanoelectronic devices from self-assembled biological templates.

DNA nanotechnology for nanophotonic applications.

PubMed

Samanta, Anirban; Banerjee, Saswata; Liu, Yan

2015-02-14

DNA nanotechnology has touched the epitome of miniaturization by integrating various nanometer size particles with nanometer precision. This enticing bottom-up approach has employed small DNA tiles, large multi-dimensional polymeric structures or more recently DNA origami to organize nanoparticles of different inorganic materials, small organic molecules or macro-biomolecules like proteins, and RNAs into fascinating patterns that are difficult to achieve by other conventional methods. Here, we are especially interested in the self-assembly of nanomaterials that are potentially attractive elements in the burgeoning field of nanophotonics. These materials include plasmonic nanoparticles, quantum dots, fluorescent organic dyes, etc. DNA based self-assembly allows excellent control over distance, orientation and stoichiometry of these nano-elements that helps to engineer intelligent systems that can potentially pave the path for future technology. Many outstanding structures have been fabricated that are capable of fine tuning optical properties, such as fluorescence intensity and lifetime modulation, enhancement of Raman scattering and emergence of circular dichroism responses. Within the limited scope of this review we have tried to give a glimpse of the development of this still nascent but highly promising field to its current status as well as the existing challenges before us.

Tailoring charge density and hydrogen bonding of imidazolium copolymers for efficient gene delivery.

PubMed

Allen, Michael H; Green, Matthew D; Getaneh, Hiwote K; Miller, Kevin M; Long, Timothy E

2011-06-13

Conventional free radical polymerization with subsequent postpolymerization modification afforded imidazolium copolymers with controlled charge density and side chain hydroxyl number. Novel imidazolium-containing copolymers where each permanent cation contained one or two adjacent hydroxyls allowed precise structure-transfection efficiency studies. The degree of polymerization was identical for all copolymers to eliminate the influence of molecular weight on transfection efficiency. DNA binding, cytotoxicity, and in vitro gene transfection in African green monkey COS-7 cells revealed structure-property-transfection relationships for the copolymers. DNA gel shift assays indicated that higher charge densities and hydroxyl concentrations increased DNA binding. As the charge density of the copolymers increased, toxicity of the copolymers also increased; however, as hydroxyl concentration increased, cytotoxicity remained constant. Changing both charge density and hydroxyl levels in a systematic fashion revealed a dramatic influence on transfection efficiency. Dynamic light scattering of the polyplexes, which were composed of copolymer concentrations required for the highest luciferase expression, showed an intermediate DNA-copolymer binding affinity. Our studies supported the conclusion that cationic copolymer binding affinity significantly impacts overall transfection efficiency of DNA delivery vehicles, and the incorporation of hydroxyl sites offers a less toxic and effective alternative to more conventional highly charged copolymers.

Complementary molecular information changes our perception of food web structure

PubMed Central

Wirta, Helena K.; Hebert, Paul D. N.; Kaartinen, Riikka; Prosser, Sean W.; Várkonyi, Gergely; Roslin, Tomas

2014-01-01

How networks of ecological interactions are structured has a major impact on their functioning. However, accurately resolving both the nodes of the webs and the links between them is fraught with difficulties. We ask whether the new resolution conferred by molecular information changes perceptions of network structure. To probe a network of antagonistic interactions in the High Arctic, we use two complementary sources of molecular data: parasitoid DNA sequenced from the tissues of their hosts and host DNA sequenced from the gut of adult parasitoids. The information added by molecular analysis radically changes the properties of interaction structure. Overall, three times as many interaction types were revealed by combining molecular information from parasitoids and hosts with rearing data, versus rearing data alone. At the species level, our results alter the perceived host specificity of parasitoids, the parasitoid load of host species, and the web-wide role of predators with a cryptic lifestyle. As the northernmost network of host–parasitoid interactions quantified, our data point exerts high leverage on global comparisons of food web structure. However, how we view its structure will depend on what information we use: compared with variation among networks quantified at other sites, the properties of our web vary as much or much more depending on the techniques used to reconstruct it. We thus urge ecologists to combine multiple pieces of evidence in assessing the structure of interaction webs, and suggest that current perceptions of interaction structure may be strongly affected by the methods used to construct them. PMID:24449902

One-Dimensional Multichromophor Arrays Based on DNA: From Self-Assembly to Light-Harvesting.

PubMed

Ensslen, Philipp; Wagenknecht, Hans-Achim

2015-10-20

Light-harvesting complexes collect light energy and deliver it by a cascade of energy and electron transfer processes to the reaction center where charge separation leads to storage as chemical energy. The design of artificial light-harvesting assemblies faces enormous challenges because several antenna chromophores need to be kept in close proximity but self-quenching needs to be avoided. Double stranded DNA as a supramolecular scaffold plays a promising role due to its characteristic structural properties. Automated DNA synthesis allows incorporation of artificial chromophore-modified building blocks, and sequence design allows precise control of the distances and orientations between the chromophores. The helical twist between the chromophores, which is induced by the DNA framework, controls energy and electron transfer and thereby reduces the self-quenching that is typically observed in chromophore aggregates. This Account summarizes covalently multichromophore-modified DNA and describes how such multichromophore arrays were achieved by Watson-Crick-specific and DNA-templated self-assembly. The covalent DNA systems were prepared by incorporation of chromophores as DNA base substitutions (either as C-nucleosides or with acyclic linkers as substitutes for the 2'-deoxyribofuranoside) and as DNA base modifications. Studies with DNA base substitutions revealed that distances but more importantly relative orientations of the chromophores govern the energy transfer efficiencies and thereby the light-harvesting properties. With DNA base substitutions, duplex stabilization was faced and could be overcome, for instance, by zipper-like placement of the chromophores in both strands. For both principal structural approaches, DNA-based light-harvesting antenna could be realized. The major disadvantages, however, for covalent multichromophore DNA conjugates are the poor yields of synthesis and the solubility issues for oligonucleotides with more than 5-10 chromophore modifications in a row. A logical alternative approach is to leave out the phosphodiester bridges between the chromophores and let chromophore-nucleoside conjugates self-assemble specifically along single stranded DNA as template. The self-organization of chromophores along the DNA template based on canonical base pairing would be advantageous because sequence selective base pairing could provide a structural basis for programmed complexity within the chromophore assembly. The self-assembly is governed by two interactions. The chromophore-nucleoside conjugates as guest molecules are recognized via hydrogen bonds to the corresponding counter bases in the single stranded DNA template. Moreover, the π-π interactions between the stacked chromophores stabilize these self-assembled constructs with increasing length. Longer DNA templates are more attractive for self-assembled antenna. The helicity in the stack of porphyrins as guest molecules assembled on the DNA template can be switched by environmental changes, such as pH variations. DNA-templated stacks of ethynyl pyrene and nile red exhibit left-handed chirality, which stands in contrast to similar covalent multichromophore-DNA conjugates with enforced right-handed helicity. With ethynyl nile red, it is possible to occupy every available binding site on the templates. Mixed assemblies of ethynyl pyrene and nile red show energy transfer and thereby provide a proof-of-principle that simple light-harvesting antennae can be obtained in a noncovalent and self-assembled fashion. With respect to the next important step, chemical storage of the absorbed light energy, future research has to focus on the coupling of sophisticated DNA-based light-harvesting antenna to reaction centers.

Nanoparticles and DNA - a powerful and growing functional combination in bionanotechnology

NASA Astrophysics Data System (ADS)

Samanta, Anirban; Medintz, Igor L.

2016-04-01

Functionally integrating DNA and other nucleic acids with nanoparticles in all their different physicochemical forms has produced a rich variety of composite nanomaterials which, in many cases, display unique or augmented properties due to the synergistic activity of both components. These capabilities, in turn, are attracting greater attention from various research communities in search of new nanoscale tools for diverse applications that include (bio)sensing, labeling, targeted imaging, cellular delivery, diagnostics, therapeutics, theranostics, bioelectronics, and biocomputing to name just a few amongst many others. Here, we review this vibrant and growing research area from the perspective of the materials themselves and their unique capabilities. Inorganic nanocrystals such as quantum dots or those made from gold or other (noble) metals along with metal oxides and carbon allotropes are desired as participants in these hybrid materials since they can provide distinctive optical, physical, magnetic, and electrochemical properties. Beyond this, synthetic polymer-based and proteinaceous or viral nanoparticulate materials are also useful in the same role since they can provide a predefined and biocompatible cargo-carrying and targeting capability. The DNA component typically provides sequence-based addressability for probes along with, more recently, unique architectural properties that directly originate from the burgeoning structural DNA field. Additionally, DNA aptamers can also provide specific recognition capabilities against many diverse non-nucleic acid targets across a range of size scales from ions to full protein and cells. In addition to appending DNA to inorganic or polymeric nanoparticles, purely DNA-based nanoparticles have recently surfaced as an excellent assembly platform and have started finding application in areas like sensing, imaging and immunotherapy. We focus on selected and representative nanoparticle-DNA materials and highlight their myriad applications using examples from the literature. Overall, it is clear that this unique functional combination of nanomaterials has far more to offer than what we have seen to date and as new capabilities for each of these materials are developed, so, too, will new applications emerge.

DNA condensing effects and sequence selectivity of DNA binding of antitumor noncovalent polynuclear platinum complexes.

PubMed

Malina, Jaroslav; Farrell, Nicholas P; Brabec, Viktor

2014-02-03

The noncovalent analogues of antitumor polynuclear platinum complexes represent a structurally discrete class of platinum drugs. Their chemical and biological properties differ significantly from those of most platinum chemotherapeutics, which bind to DNA in a covalent manner by formation of Pt-DNA adducts. In spite of the fact that these noncovalent polynuclear platinum complexes contain no leaving groups, they have been shown to bind to DNA with high affinity. We report here on the DNA condensation properties of a series of noncovalent analogues of antitumor polynuclear platinum complexes described by biophysical and biochemical methods. The results demonstrate that these polynuclear platinum compounds are capable of inducing DNA condensation at more than 1 order of magnitude lower concentrations than conventional spermine. Atomic force microscopy studies of DNA condensation confined to a mica substrate have revealed that the DNA morphologies become more compact with increasing concentration of the platinum complexes. Moreover, we also found that the noncovalent polynuclear platinum complex [{Pt(NH3)3}2-μ-{trans-Pt(NH3)2(NH2(CH2)6NH2)2}](6+) (TriplatinNC-A) binds to DNA in a sequence-dependent manner, namely, to A/T-rich sequences and A-tract regions, and that noncovalent polynuclear platinum complexes protect DNA from enzymatic cleavage by DNase I. The results suggest that mechanisms of antitumor and cytotoxic activities of these complexes may be associated with their unique ability to condense DNA along with their sequence-specific DNA binding. Owing to their high cellular accumulation, it is also reasonable to suggest that their mechanism of action is based on the competition with naturally occurring DNA condensing agents, such as polyamines spermine, spermidine, and putrescine, for intracellular binding sites, resulting in the disturbance of the correct binding of regulatory proteins initiating the onset of apoptosis.

Nanoscale superstructures assembled by polymerase chain reaction (PCR): programmable construction, structural diversity, and emerging applications.

PubMed

Kuang, Hua; Ma, Wei; Xu, Liguang; Wang, Libing; Xu, Chuanlai

2013-11-19

Polymerase chain reaction (PCR) is an essential tool in biotechnology laboratories and is becoming increasingly important in other areas of research. Extensive data obtained over the last 12 years has shown that the combination of PCR with nanoscale dispersions can resolve issues in the preparation DNA-based materials that include both inorganic and organic nanoscale components. Unlike conventional DNA hybridization and antibody-antigen complexes, PCR provides a new, effective assembly platform that both increases the yield of DNA-based nanomaterials and allows researchers to program and control assembly with predesigned parameters including those assisted and automated by computers. As a result, this method allows researchers to optimize to the combinatorial selection of the DNA strands for their nanoparticle conjugates. We have developed a PCR approach for producing various nanoscale assemblies including organic motifs such as small molecules, macromolecules, and inorganic building blocks, such as nanorods (NRs), metal, semiconductor, and magnetic nanoparticles (NPs). We start with a nanoscale primer and then modify that building block using the automated steps of PCR-based assembly including initialization, denaturation, annealing, extension, final elongation, and final hold. The intermediate steps of denaturation, annealing, and extension are cyclic, and we use computer control so that the assembled superstructures reach their predetermined complexity. The structures assembled using a small number of PCR cycles show a lower polydispersity than similar discrete structures obtained by direct hybridization between the nanoscale building blocks. Using different building blocks, we assembled the following structural motifs by PCR: (1) discrete nanostructures (NP dimers, NP multimers including trimers, pyramids, tetramers or hexamers, etc.), (2) branched NP superstructures and heterochains, (3) NP satellite-like superstructures, (4) Y-shaped nanostructures and DNA networks, (5) protein-DNA co-assembly structures, and (6) DNA block copolymers including trimers and pentamers. These results affirm that this method can produce a variety of chemical structures and in yields that are tunable. Using PCR-based preparation of DNA-bridged nanostructures, we can program the assembly of the nanoscale blocks through the adjustment of the primer intensity on the assembled units, the number of PCR cycles, or both. The resulting structures are highly complex and diverse and have interesting dynamics and collective properties. Potential applications of these materials include chirooptical materials, probe fabrication, and environmental and biomedical sensors.

Synthesis, crystal structure, DFT calculation and DNA binding studies of new water-soluble derivatives of dppz

NASA Astrophysics Data System (ADS)

Aminzadeh, Mohammad; Eslami, Abbas; Kia, Reza; Aleeshah, Roghayeh

2017-10-01

Diquaternarization of dipyrido-[2,3-a:2‧,3‧-c]-phenazine,(dppz) and its analogous dipyrido-[2,3-a:2‧,3‧-c]-dimethylphenazine,(dppx) using 1,3-dibromopropane afford new water-soluble derivatives of phenazine, propylene-bipyridyldiylium-phenazine (1) and propylene-bipyridyldiylium-dimethylphenazine (2). The compounds have been characterized by means of FT-IR, NMR, elemental analysis and conductometric measurements and their structure were determined by X-ray crystallography. The experimental studies on the compounds have been accompanied computationally by Density Functional Theory (DFT) calculations. The DNA binding properties of both compounds to calf thymus DNA (ctDNA) were investigated by UV-Vis absorption and emission methods. The expanded UV-Vis spectral data matrix was analyzed by multivariate curve resolution-alternating least squares (MCR-ALS) technique to obtain the concentration profile and pure spectra of all reaction species which existed in the interaction procedure. Multivariate curve resolution may help us to give a better understanding of the 1(Cl)2-ctDNA and 2(Cl)2-ctDNA interaction mechanism. The results suggest that both compounds bind tightly to DNA through intercalation mechanism and the DNA binding affinity of 2 is slightly lower than that of 1 due to steric hindrance of the methyl group. Also, thermal denaturation studies reveal that these compounds show strong affinity for binding with calf thymus DNA. The thermodynamic parameters of the DNA binding process were obtained from the temperature dependence of the binding constants and the results showed that binding of both compounds to DNA is an enthalpically driven process that is in agreement with proposed DNA intercalation capability of these compounds.

Fish DNA-modified clays: Towards highly flame retardant polymer nanocomposite with improved interfacial and mechanical performance.

PubMed

Zabihi, Omid; Ahmadi, Mojtaba; Khayyam, Hamid; Naebe, Minoo

2016-12-05

Deoxyribonucleic Acid (DNA) has been recently found to be an efficient renewable and environmentally-friendly flame retardant. In this work, for the first time, we have used waste DNA from fishing industry to modify clay structure in order to increase the clay interactions with epoxy resin and take benefit of its additional thermal property effect on thermo-physical properties of epoxy-clay nanocomposites. Intercalation of DNA within the clay layers was accomplished in a one-step approach confirmed by FT-IR, XPS, TGA, and XRD analyses, indicating that d-space of clay layers was expanded from ~1.2 nm for pristine clay to ~1.9 nm for clay modified with DNA (d-clay). Compared to epoxy nanocomposite containing 2.5%wt of Nanomer I.28E organoclay (m-clay), it was found that at 2.5%wt d-clay loading, significant enhancements of ~14%, ~6% and ~26% in tensile strength, tensile modulus, and fracture toughness of epoxy nanocomposite can be achieved, respectively. Effect of DNA as clay modifier on thermal performance of epoxy nanocomposite containing 2.5%wt d-clay was evaluated using TGA and cone calorimetry analysis, revealing significant decreases of ~4000 kJ/m 2 and ~78 kW/m 2 in total heat release and peak of heat release rate, respectively, in comparison to that containing 2.5%wt of m-clay.

Fish DNA-modified clays: Towards highly flame retardant polymer nanocomposite with improved interfacial and mechanical performance

NASA Astrophysics Data System (ADS)

Zabihi, Omid; Ahmadi, Mojtaba; Khayyam, Hamid; Naebe, Minoo

2016-12-01

Deoxyribonucleic Acid (DNA) has been recently found to be an efficient renewable and environmentally-friendly flame retardant. In this work, for the first time, we have used waste DNA from fishing industry to modify clay structure in order to increase the clay interactions with epoxy resin and take benefit of its additional thermal property effect on thermo-physical properties of epoxy-clay nanocomposites. Intercalation of DNA within the clay layers was accomplished in a one-step approach confirmed by FT-IR, XPS, TGA, and XRD analyses, indicating that d-space of clay layers was expanded from ~1.2 nm for pristine clay to ~1.9 nm for clay modified with DNA (d-clay). Compared to epoxy nanocomposite containing 2.5%wt of Nanomer I.28E organoclay (m-clay), it was found that at 2.5%wt d-clay loading, significant enhancements of ~14%, ~6% and ~26% in tensile strength, tensile modulus, and fracture toughness of epoxy nanocomposite can be achieved, respectively. Effect of DNA as clay modifier on thermal performance of epoxy nanocomposite containing 2.5%wt d-clay was evaluated using TGA and cone calorimetry analysis, revealing significant decreases of ~4000 kJ/m2 and ~78 kW/m2 in total heat release and peak of heat release rate, respectively, in comparison to that containing 2.5%wt of m-clay.

Fish DNA-modified clays: Towards highly flame retardant polymer nanocomposite with improved interfacial and mechanical performance

PubMed Central

Zabihi, Omid; Ahmadi, Mojtaba; Khayyam, Hamid; Naebe, Minoo

2016-01-01

Deoxyribonucleic Acid (DNA) has been recently found to be an efficient renewable and environmentally-friendly flame retardant. In this work, for the first time, we have used waste DNA from fishing industry to modify clay structure in order to increase the clay interactions with epoxy resin and take benefit of its additional thermal property effect on thermo-physical properties of epoxy-clay nanocomposites. Intercalation of DNA within the clay layers was accomplished in a one-step approach confirmed by FT-IR, XPS, TGA, and XRD analyses, indicating that d-space of clay layers was expanded from ~1.2 nm for pristine clay to ~1.9 nm for clay modified with DNA (d-clay). Compared to epoxy nanocomposite containing 2.5%wt of Nanomer I.28E organoclay (m-clay), it was found that at 2.5%wt d-clay loading, significant enhancements of ~14%, ~6% and ~26% in tensile strength, tensile modulus, and fracture toughness of epoxy nanocomposite can be achieved, respectively. Effect of DNA as clay modifier on thermal performance of epoxy nanocomposite containing 2.5%wt d-clay was evaluated using TGA and cone calorimetry analysis, revealing significant decreases of ~4000 kJ/m2 and ~78 kW/m2 in total heat release and peak of heat release rate, respectively, in comparison to that containing 2.5%wt of m-clay. PMID:27917901

Release of DNA from polyelectrolyte multilayers fabricated using 'charge-shifting' cationic polymers: tunable temporal control and sequential, multi-agent release.

PubMed

Sun, Bin; Lynn, David M

2010-11-20

We report an approach to the design of multilayered polyelectrolyte thin films (or 'polyelectrolyte multilayers', PEMs) that can be used to provide tunable control over the release of plasmid DNA (or multiple different DNA constructs) from film-coated surfaces. Our approach is based upon methods for the layer-by-layer assembly of DNA-containing thin films, and exploits the properties of a new class of cationic 'charge-shifting' polymers (amine functionalized polymers that undergo gradual changes in net charge upon side chain ester hydrolysis) to provide control over the rates at which these films erode and release DNA. We synthesized two 'charge-shifting' polymers (polymers 1 and 2) containing different side chain structures by ring-opening reactions of poly(2-alkenyl azlactone)s with two different tertiary amine functionalized alcohols (3-dimethylamino-1-propanol and 2-dimethylaminoethanol, respectively). Subsequent characterization revealed large changes in the rates of side chain ester hydrolysis for these two polymers; whereas the half-life for the hydrolysis of the esters in polymer 1 was ~200 days, the half-life for polymer 2 was ~6 days. We demonstrate that these large differences in side chain hydrolysis make possible the design of PEMs that erode and promote the surface-mediated release of DNA either rapidly (e.g., over ~3 days for films fabricated using polymer 2) or slowly (e.g., over ~1 month for films fabricated using polymer 1). We demonstrate further that it is possible to design films with release profiles that are intermediate to these two extremes by fabricating films using solutions containing different mixtures of these two polymers. This approach can thus expand the usefulness of these two polymers and achieve a broader range of DNA release profiles without the need to synthesize polymers with new structures or properties. Finally, we demonstrate that polymers 1 and 2 can be used to fabricate multilayered films with hierarchical structures that promote the sequential release of two different DNA constructs with separate and distinct release profiles (e.g., the release of a first construct over a period of ~3 days, followed by the sustained release of a second for a period of ~70 days). With further development, this approach could contribute to the design of functional thin films and surface coatings that provide sophisticated control over the timing and the order of the release of two or more DNA constructs (or other agents) of interest in a range of biomedical contexts. Copyright © 2010 Elsevier B.V. All rights reserved.

Monodisperse Multidimensional Nanostructures via Centrifugal Separation

NASA Astrophysics Data System (ADS)

Shin, Yu Jin

Anisotropic nanomaterials, including zero-dimensional metallic nanoparticles (MNPs), one-dimensional single-walled carbon nanotubes (SWCNTs), and two-dimensional few-layer black phosphorous (FL-P) exhibit interesting structure-dependent properties that could be exploited in biomedicine, plasmonics, and optoelectronics. In this thesis, centrifugation sorting of these nanomaterials is utilized for structure refinement, investigation of structure-dependent optical response, and applications in biomedical imaging and plasmonics. Nobel NMPs show unique shape- and size-dependent optical properties. Controlled synthetic methods are developed to manipulate the structure of these NMPs, but intrinsically produce dispersions of polydisperse NPs with various shape and size, and synthetic byproducts. Here, we describe a facile strategy for separating small (edge length <100 nm) faceted gold NPs: rhombic dodecahedra (RD) and obtuse triangular bipyramids (BPs), which form simultaneously during synthesis but are hard to separate via commercial filters. By utilizing centrifugation of the as-synthesized mixture in a shallow density gradient centrifugation (DGC), we are able to isolate a high purity of BPs (>80%) and subsequently achieve a 2.5 fold enhancement in refractive index sensitivity, comparable to the unsorted mixture. This shallow DGC approach is robust and reliable, and therefore can be applied to other metal nanostructures for concomitant improvements in plasmonic properties and applications. Using the identical separation strategy in the previous study, we are able to enrich gold nanostars as a function of branch number. In particular, we explore different variants of density gradient media to ensure compatibility with the star shape and colloid stability. We determine that sucrose is compatible with nanostars stability and surface functionalizaton. The refined population of gold stars are functionalized with Gd(III)-DNA to act as MRI contrast agents, and thus enables us to investigate how populations of nanostars with different branch numbers contribute to the relaxivity of surface bound Gd(III)-DNA. It is shown that the increased relaxivity of DNA-Gd star is correlated with increased number of star branches, not with increased size of the stars. Therefore, shape is a new parameter which can be tuned in the design of NP-based MRI contrast agent. These findings can also improve the performance of functionalized anisotropic nanoconjugates which have potential for applications such as lowering detection limits for sensors and diagnostics, or enabling new modes of self-assembly. Finally, we have broadened the scope of DGC to other dimensional nanomaterials: 1D-SWCNTs and 2D-FL-P. Despite their tunable and structure-dependent optical properties, intrinsic structural heterogeneity and poor quantum efficiency limit their potential applications. Therefore, DGC is employed to separate the SWCNTs and FL-P by length and the number of layers, respectively, thereby incorporating them into optical cavity structures for enhancing their luminescence properties. These fundamental studies of low-dimensional nanomaterials assist in the design process for optoelectronic device fabrication.

Structural Transitions in Supercoiled Stretched DNA

NASA Astrophysics Data System (ADS)

v, Croquette

1998-03-01

Using magnetic micromanipulation techniques [Strick 96]( uc(T.R.) Strick, J.-F. Allemand, D. Bensimon, A. Bensimon) and uc(V.) Croquette, "The elasticity of a single supercoiled DNA molecule", Science, 271, 1835 (1996)., we have studied the mechanical properties (force versus extension) of single DNA molecules under a wide range of torsional stresses (supercoiling). We show that unwinding the DNA double helix leads to a phase separation between regular B-DNA and denaturation bubbles. The fraction of denatured molecule increases linearly with the degree of unwinding, beginning at a value of 1% unwinding. We have confirmed this denatured state by hybridization of homologous single-stranded DNA probes and by a chemical attack of the exposed bases. Surprisingly, when we overwind the molecule, the elasticity curves we obtain may also be interpreted by the coexistence of two phases, B-DNA and a new phase which we note P-DNA. The fraction of this new phase increases smoothly with overwinding, beginning at 3 % and continuing up to 300 %. Our results indicate that this new phase is four times more twisted that the standard B-DNA and is 1.75 times longer. Although the structure of this phase is not yet known, such a high twisting can only be attained if the sugar-phosphate backbones of the two strands are twisted closely while the bases are expelled outside of the molecule's core, in a structure reminiscent of the one proposed by Pauling. Indeed we have shown that this new phase is sensitive to chemical attack whereas the B-DNA is not. This new phase begins to appear on a molecule overwound by 3 % and stretched by a force of 5 pN, conditions typically encountered in vivo during gene transcription. This new phase may thus play a biological role (for more details).

Fluorescence of size-expanded DNA bases: reporting on DNA sequence and structure with an unnatural genetic set.

PubMed

Krueger, Andrew T; Kool, Eric T

2008-03-26

We recently described the synthesis and helix assembly properties of expanded DNA (xDNA), which contains base pairs 2.4 A larger than natural DNA pairs. This designed genetic set is under study with the goals of mimicking the functions of the natural DNA-based genetic system and of developing useful research tools. Here, we study the fluorescence properties of the four expanded bases of xDNA (xA, xC, xG, xT) and evaluate how their emission varies with changes in oligomer length, composition, and hybridization. Experiments were carried out with short oligomers of xDNA nucleosides conjugated to a DNA oligonucleotide, and we investigated the effects of hybridizing these fluorescent oligomers to short complementary DNAs with varied bases opposite the xDNA bases. As monomer nucleosides, the xDNA bases absorb light in two bands: one at approximately 260 nm (similar to DNA) and one at longer wavelength ( approximately 330 nm). All are efficient violet-blue fluorophores with emission maxima at approximately 380-410 nm and quantum yields (Phifl) of 0.30-0.52. Short homo-oligomers of the xDNA bases (length 1-4 monomers) showed moderate self-quenching except xC, which showed enhancement of Phifl with increasing length. Interestingly, multimers of xA emitted at longer wavelengths (520 nm) as an apparent excimer. Hybridization of an oligonucleotide to the DNA adjacent to the xDNA bases (with the xDNA portion overhanging) resulted in no change in fluorescence. However, addition of one, two, or more DNA bases in these duplexes opposite the xDNA portion resulted in a number of significant fluorescence responses, including wavelength shifts, enhancements, or quenching. The strongest responses were the enhancement of (xG)n emission by hybridization of one or more adenines opposite them, and the quenching of (xT)n and (xC)n emission by guanines opposite. The data suggest multiple ways in which the xDNA bases, both alone and in oligomers, may be useful as tools in biophysical analysis and biotechnological applications.

Fungal beta glucan protects radiation induced DNA damage in human lymphocytes

PubMed Central

Maurya, Dharmendra K.; Salvi, Veena P.; Janardhanan, Krishnankutty K; Nair, Cherupally K. K.

2014-01-01

Background Ganoderma lucidum (Ling Zhi), a basidiomycete white rot macrofungus has been used extensively for therapeutic use in China, Japan, Korea and other Asian countries for 2,000 years. The present study is an attempt to investigate its DNA protecting property in human lymphocytes. Materials and methods Beta glucan (BG) was isolated by standard procedure and the structure and composition were studied by infrared radiation (IR) and nuclear magnetic resonance (NMR) spectroscopy, gel filtration chromatography and paper chromatography. The radioprotective properties of BG isolated from the macro fungi Ganoderma lucidum was assessed by single cell gel electrophoresis (comet assay). Human lymphocytes were exposed to 0, 1, 2 and 4 Gy gamma radiation in the presence and absence of BG. Results The comet parameters were reduced by BG. The results indicate that the BG of G. lucidum possessed significant radioprotective activity with DNA repairing ability and antioxidant activity as the suggestive mechanism. Conclusions The findings suggest the potential use of this mushroom for the prevention of radiation induced cellular damages. PMID:25332989

Fungal beta glucan protects radiation induced DNA damage in human lymphocytes.

PubMed

Pillai, Thulasi G; Maurya, Dharmendra K; Salvi, Veena P; Janardhanan, Krishnankutty K; Nair, Cherupally K K

2014-02-01

Ganoderma lucidum (Ling Zhi), a basidiomycete white rot macrofungus has been used extensively for therapeutic use in China, Japan, Korea and other Asian countries for 2,000 years. The present study is an attempt to investigate its DNA protecting property in human lymphocytes. Beta glucan (BG) was isolated by standard procedure and the structure and composition were studied by infrared radiation (IR) and nuclear magnetic resonance (NMR) spectroscopy, gel filtration chromatography and paper chromatography. The radioprotective properties of BG isolated from the macro fungi Ganoderma lucidum was assessed by single cell gel electrophoresis (comet assay). Human lymphocytes were exposed to 0, 1, 2 and 4 Gy gamma radiation in the presence and absence of BG. The comet parameters were reduced by BG. The results indicate that the BG of G. lucidum possessed significant radioprotective activity with DNA repairing ability and antioxidant activity as the suggestive mechanism. The findings suggest the potential use of this mushroom for the prevention of radiation induced cellular damages.

Synthesis, crystal structure and interaction of L-valine Schiff base divanadium(V) complex containing a V2O3 core with DNA and BSA.

PubMed

Guo, Qiong; Li, Lianzhi; Dong, Jianfang; Liu, Hongyan; Xu, Tao; Li, Jinghong

2013-04-01

A divanadium(V) complex, [V2O3(o-van-val)2] (o-van-val=Schiff base derived from o-vanillin and L-valine), has been synthesized and structurally characterized. The crystal structure shows that both of the vanadium centers in the complex have a distorted octahedral coordination environment composed of tridentate Schiff base ligand. A V2O3 core in molecular structure adopts intermediate between cis and trans configuration with the O1V1⋯V1AO1A torsion angle 115.22 (28)° and the V1⋯V1A distance 3.455Å. The binding properties of the complex with calf thymus DNA (CT-DNA) have been investigated by UV-vis absorption, fluorescence, CD spectra and viscosity measurement. The results indicate that the complex binds to CT-DNA in non-classical intercalative mode. Meanwhile, the interaction of the complex with bovine serum albumin (BSA) has been studied by UV-vis absorption, fluorescence and CD spectra. Results indicated that the complex can markedly quench the intrinsic fluorescence of BSA via a static quenching process, and cause its conformational change. The calculated apparent binding constant Kb was 1.05×10(6)M(-1) and the binding site number n was 1.18. Copyright © 2013 Elsevier B.V. All rights reserved.

High-resolution biophysical analysis of the dynamics of nucleosome formation

PubMed Central

Hatakeyama, Akiko; Hartmann, Brigitte; Travers, Andrew; Nogues, Claude; Buckle, Malcolm

2016-01-01

We describe a biophysical approach that enables changes in the structure of DNA to be followed during nucleosome formation in in vitro reconstitution with either the canonical “Widom” sequence or a judiciously mutated sequence. The rapid non-perturbing photochemical analysis presented here provides ‘snapshots’ of the DNA configuration at any given moment in time during nucleosome formation under a very broad range of reaction conditions. Changes in DNA photochemical reactivity upon protein binding are interpreted as being mainly induced by alterations in individual base pair roll angles. The results strengthen the importance of the role of an initial (H3/H4)2 histone tetramer-DNA interaction and highlight the modulation of this early event by the DNA sequence. (H3/H4)2 binding precedes and dictates subsequent H2A/H2B-DNA interactions, which are less affected by the DNA sequence, leading to the final octameric nucleosome. Overall, our results provide a novel, exciting way to investigate those biophysical properties of DNA that constitute a crucial component in nucleosome formation and stabilization. PMID:27263658

Identification of the DNA-Binding Domains of Human Replication Protein A That Recognize G-Quadruplex DNA

PubMed Central

Prakash, Aishwarya; Natarajan, Amarnath; Marky, Luis A.; Ouellette, Michel M.; Borgstahl, Gloria E. O.

2011-01-01

Replication protein A (RPA), a key player in DNA metabolism, has 6 single-stranded DNA-(ssDNA-) binding domains (DBDs) A-F. SELEX experiments with the DBDs-C, -D, and -E retrieve a 20-nt G-quadruplex forming sequence. Binding studies show that RPA-DE binds preferentially to the G-quadruplex DNA, a unique preference not observed with other RPA constructs. Circular dichroism experiments show that RPA-CDE-core can unfold the G-quadruplex while RPA-DE stabilizes it. Binding studies show that RPA-C binds pyrimidine- and purine-rich sequences similarly. This difference between RPA-C and RPA-DE binding was also indicated by the inability of RPA-CDE-core to unfold an oligonucleotide containing a TC-region 5′ to the G-quadruplex. Molecular modeling studies of RPA-DE and telomere-binding proteins Pot1 and Stn1 reveal structural similarities between the proteins and illuminate potential DNA-binding sites for RPA-DE and Stn1. These data indicate that DBDs of RPA have different ssDNA recognition properties. PMID:21772997

Comparison of Iron-Binding Ability Between Thr70-NapA and Ser70-NapA of Helicobacter pylori.

PubMed

Shan, Weiran; Kung, Hsiang-Fu; Ge, Ruiguang

2016-06-01

The neutrophil-activating protein (NapA) of Helicobacter pylori (H. pylori), with DNA-binding and iron seizing properties, is a fundamental virulence factor involved in H. pylori-related diseases. Compared with Ser70-NapA strain, Thr70-NapA strain is more intimately correlated with iron-deficiency anemia. To investigate whether two types of proteins differ in iron-binding ability, mutated Thr70-NapA and Ser70-NapA strains were established. Isothermal titration calorimetry (ITC) method was conducted to measure the binding between the NapA protein and Fe(2+) . The structural changes of NapA protein were also tested during iron interaction by fast protein liquid chromatography (FPLC) and circular dichroism (CD) methods. DNA-binding assay was performed for evaluate the affinity of both mutated and wild types of NapA with DNA. Mutated Thr70-NapA had higher iron-binding ability than wild Ser70-NapA. The structural stability of Thr70-NapA was disrupted and became more active along with the rising concentration of Fe(2+) , whereas no similar association was observed between Ser70-NapA and Fe(2+) level. When the iron/protein molar ratio ranged from 10 to 20, both Ser70-NapA and Thr70-NapA displayed weaker DNA-binding ability. Thr70-NapA has much stronger ability to sequester ferrous ion compared with Ser70-NapA in H. pylori. In addition, the DNA-binding property of NapA is dependent upon the Fe(2+) concentration. © 2015 John Wiley & Sons Ltd.

Molecular mechanical studies of DNA flexibility: Coupled backbone torsion angles and base-pair openings

PubMed Central

Keepers, Joe W.; Kollman, Peter A.; Weiner, Paul K.; James, Thomas L.

1982-01-01

Molecular mechanics studies have been carried out on “B-DNA-like” structures of [d(C-G-C-G-A-A-T-T-C-G-C-G)]2 and [d(A)]12·[d(T)]12. Each of the backbone torsion angles (ψ, φ, ω, ω′, φ′) has been “forced” to alternative values from the normal B-DNA values (g+, t, g-, g-, t conformations). Compensating torsion angle changes preserve most of the base stacking energy in the double helix. In a second part of the study, one purine N3-pyrimidine N1 distance at a time has been forced to a value of 6 Å in an attempt to simulate the base opening motions required to rationalize proton exchange data for DNA. When the 6-Å constraint is removed, many of the structures revert to the normal Watson-Crick hydrogen-bonded structure, but a number are trapped in structures ≈5 kcal/mol higher in energy than the starting B-DNA structure. The relative energy of these structures, some of which involve a non-Watson-Crick thymine C2(carbonyl)[unk]adenine 6NH2 hydrogen bond, are qualitatively consistent with the ΔH for a “base pair-open state” suggested by Mandal et al. of 4-6 kcal/mol [Mandal, C., Kallenbach, N. R. & Englander, S. W. (1979) J. Mol. Biol. 135, 391-411]. The picture of DNA flexibility emerging from this study depicts the backbone as undergoing rapid motion between local torsional minima on a nanosecond time scale. Backbone motion is mainly localized within a dinucleoside segment and generally not conformationally coupled along the chain or across the base pairs. Base motions are much smaller in magnitude than backbone motions. Base sliding allows imino N—H exchange, but it is localized, and only a small fraction of the N—H groups is exposed at any one time. Stacking and hydrogen bonding cause a rigid core of bases in the center of the molecule accounting for the hydrodynamic properties of DNA. PMID:6957879

DNA "nano-claw": logic-based autonomous cancer targeting and therapy.

PubMed

You, Mingxu; Peng, Lu; Shao, Na; Zhang, Liqin; Qiu, Liping; Cui, Cheng; Tan, Weihong

2014-01-29

Cell types, both healthy and diseased, can be classified by inventories of their cell-surface markers. Programmable analysis of multiple markers would enable clinicians to develop a comprehensive disease profile, leading to more accurate diagnosis and intervention. As a first step to accomplish this, we have designed a DNA-based device, called "Nano-Claw". Combining the special structure-switching properties of DNA aptamers with toehold-mediated strand displacement reactions, this claw is capable of performing autonomous logic-based analysis of multiple cancer cell-surface markers and, in response, producing a diagnostic signal and/or targeted photodynamic therapy. We anticipate that this design can be widely applied in facilitating basic biomedical research, accurate disease diagnosis, and effective therapy.

The architectonics of programmable RNA and DNA nanostructures.

PubMed

Jaeger, Luc; Chworos, Arkadiusz

2006-08-01

The past several years have witnessed the emergence of a new world of nucleic-acid-based architectures with highly predictable and programmable self-assembly properties. For almost two decades, DNA has been the primary material for nucleic acid nanoconstruction. More recently, the dramatic increase in RNA structural information led to the development of RNA architectonics, the scientific study of the principles of RNA architecture with the aim of constructing RNA nanostructures of any arbitrary size and shape. The remarkable modularity and the distinct but complementary nature of RNA and DNA nanomaterials are revealed by the various self-assembly strategies that aim to achieve control of the arrangement of matter at a nanoscale level.

Casting inorganic structures with DNA molds.

PubMed

Sun, Wei; Boulais, Etienne; Hakobyan, Yera; Wang, Wei Li; Guan, Amy; Bathe, Mark; Yin, Peng

2014-11-07

We report a general strategy for designing and synthesizing inorganic nanostructures with arbitrarily prescribed three-dimensional shapes. Computationally designed DNA strands self-assemble into a stiff "nanomold" that contains a user-specified three-dimensional cavity and encloses a nucleating gold "seed." Under mild conditions, this seed grows into a larger cast structure that fills and thus replicates the cavity. We synthesized a variety of nanoparticles with 3-nanometer resolution: three distinct silver cuboids with three independently tunable dimensions, silver and gold nanoparticles with diverse cross sections, and composite structures with homo- and heterogeneous components. The designer equilateral silver triangular and spherical nanoparticles exhibited plasmonic properties consistent with electromagnetism-based simulations. Our framework is generalizable to more complex geometries and diverse inorganic materials, offering a range of applications in biosensing, photonics, and nanoelectronics. Copyright © 2014, American Association for the Advancement of Science.

A Novel Charged Medium Consisting of Gas-Liquid Interfacial Plasmas

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kaneko, Toshiro; Hatakeyama, Rikizo

Due to the unique properties of ionic liquids such as their extremely low vapor pressure and high heat capacity, we succeed in creating the reactive gas (plasmas)--liquid (ionic liquids) interfacial field under a low gas pressure condition, where the plasma ion behavior can be controlled. The effects of the plasma ion irradiation on the liquid medium are quantitatively revealed for the first time. In connection with the plasma ion irradiation, the potential structure and optical emission properties of the gas-liquid interfacial plasma are investigated by changing a polarity of the electrode in the liquid to evaluate the plasma-liquid interactions. Thesemore » results would contribute to synthesizing the metal nanoparticles with carbon nanotubes as a template in the ionic liquid. It is found that the high density, mono-dispersed, and isolated metal nanoparticles are synthesized between or inside the carbon nanotubes by controlling the gas-liquid interfacial plasmas. Furthermore, we can form novel nano-bio composite materials, such as DNA encapsulated carbon nanotubes using the plasma ion irradiation method in an electrolyte plasma with DNA, and demonstrate modifications of the electrical properties of the carbon nanotubes depending on the kinds of encapsulated DNA for the first time.« less

Enzyme-guided DNA Sewing Architecture

PubMed Central

Song, In Hyun; Shin, Seung Won; Park, Kyung Soo; Lansac, Yves; Jang, Yun Hee; Um, Soong Ho

2015-01-01

With the advent of nanotechnology, a variety of nanoarchitectures with varied physicochemical properties have been designed. Owing to the unique characteristics, DNAs have been used as a functional building block for novel nanoarchitecture. In particular, a self-assembly of long DNA molecules via a piece DNA staple has been utilized to attain such constructs. However, it needs many talented prerequisites (e.g., complicated computer program) with fewer yields of products. In addition, it has many limitations to overcome: for instance, (i) thermal instability under moderate environments and (ii) restraint in size caused by the restricted length of scaffold strands. Alternatively, the enzymatic sewing linkage of short DNA blocks is simply designed into long DNA assemblies but it is more error-prone due to the undeveloped sequence data. Here, we present, for the first time, a comprehensive study for directly combining DNA structures into higher DNA sewing constructs through the 5′-end cohesive ligation of T4 enzyme. Inspired by these achievements, the synthesized DNA nanomaterials were also utilized for effective detection and real-time diagnosis of cancer-specific and cytosolic RNA markers. This generalized protocol for generic DNA sewing is expected to be useful in several DNA nanotechnology as well as any nucleic acid-related fields. PMID:26634810

DNA replication after mutagenic treatment in Hordeum vulgare.

PubMed

Kwasniewska, Jolanta; Kus, Arita; Swoboda, Monika; Braszewska-Zalewska, Agnieszka

2016-12-01

The temporal and spatial properties of DNA replication in plants related to DNA damage and mutagenesis is poorly understood. Experiments were carried out to explore the relationships between DNA replication, chromatin structure and DNA damage in nuclei from barley root tips. We quantitavely analysed the topological organisation of replication foci using pulse EdU labelling during the S phase and its relationship with the DNA damage induced by mutagenic treatment with maleic hydrazide (MH), nitroso-N-methyl-urea (MNU) and gamma ray. Treatment with mutagens did not change the characteristic S-phase patterns in the nuclei; however, the frequencies of the S-phase-labelled cells after treatment differed from those observed in the control cells. The analyses of DNA replication in barley nuclei were extended to the micronuclei induced by mutagens. Replication in the chromatin of the micronuclei was rare. The results of simultanous TUNEL reaction to identify cells with DNA strand breaks and the labelling of the S-phase cells with EdU revealed the possibility of DNA replication occurring in damaged nuclei. For the first time, the intensity of EdU fluorescence to study the rate of DNA replication was analysed. Copyright © 2016 Elsevier B.V. All rights reserved.

Enzyme-guided DNA Sewing Architecture

NASA Astrophysics Data System (ADS)

Song, In Hyun; Shin, Seung Won; Park, Kyung Soo; Lansac, Yves; Jang, Yun Hee; Um, Soong Ho

2015-12-01

With the advent of nanotechnology, a variety of nanoarchitectures with varied physicochemical properties have been designed. Owing to the unique characteristics, DNAs have been used as a functional building block for novel nanoarchitecture. In particular, a self-assembly of long DNA molecules via a piece DNA staple has been utilized to attain such constructs. However, it needs many talented prerequisites (e.g., complicated computer program) with fewer yields of products. In addition, it has many limitations to overcome: for instance, (i) thermal instability under moderate environments and (ii) restraint in size caused by the restricted length of scaffold strands. Alternatively, the enzymatic sewing linkage of short DNA blocks is simply designed into long DNA assemblies but it is more error-prone due to the undeveloped sequence data. Here, we present, for the first time, a comprehensive study for directly combining DNA structures into higher DNA sewing constructs through the 5‧-end cohesive ligation of T4 enzyme. Inspired by these achievements, the synthesized DNA nanomaterials were also utilized for effective detection and real-time diagnosis of cancer-specific and cytosolic RNA markers. This generalized protocol for generic DNA sewing is expected to be useful in several DNA nanotechnology as well as any nucleic acid-related fields.

Imparting the unique properties of DNA into complex material architectures and functions.

PubMed

Xu, Phyllis F; Noh, Hyunwoo; Lee, Ju Hun; Domaille, Dylan W; Nakatsuka, Matthew A; Goodwin, Andrew P; Cha, Jennifer N

2013-07-01

While the remarkable chemical and biological properties of DNA have been known for decades, these properties have only been imparted into materials with unprecedented function much more recently. The inimitable ability of DNA to form programmable, complex assemblies through stable, specific, and reversible molecular recognition has allowed the creation of new materials through DNA's ability to control a material's architecture and properties. In this review we discuss recent progress in how DNA has brought unmatched function to materials, focusing specifically on new advances in delivery agents, devices, and sensors.

DNA/RNA hybrid substrates modulate the catalytic activity of purified AID.

PubMed

Abdouni, Hala S; King, Justin J; Ghorbani, Atefeh; Fifield, Heather; Berghuis, Lesley; Larijani, Mani

2018-01-01

Activation-induced cytidine deaminase (AID) converts cytidine to uridine at Immunoglobulin (Ig) loci, initiating somatic hypermutation and class switching of antibodies. In vitro, AID acts on single stranded DNA (ssDNA), but neither double-stranded DNA (dsDNA) oligonucleotides nor RNA, and it is believed that transcription is the in vivo generator of ssDNA targeted by AID. It is also known that the Ig loci, particularly the switch (S) regions targeted by AID are rich in transcription-generated DNA/RNA hybrids. Here, we examined the binding and catalytic behavior of purified AID on DNA/RNA hybrid substrates bearing either random sequences or GC-rich sequences simulating Ig S regions. If substrates were made up of a random sequence, AID preferred substrates composed entirely of DNA over DNA/RNA hybrids. In contrast, if substrates were composed of S region sequences, AID preferred to mutate DNA/RNA hybrids over substrates composed entirely of DNA. Accordingly, AID exhibited a significantly higher affinity for binding DNA/RNA hybrid substrates composed specifically of S region sequences, than any other substrates composed of DNA. Thus, in the absence of any other cellular processes or factors, AID itself favors binding and mutating DNA/RNA hybrids composed of S region sequences. AID:DNA/RNA complex formation and supporting mutational analyses suggest that recognition of DNA/RNA hybrids is an inherent structural property of AID. Copyright © 2017 Elsevier Ltd. All rights reserved.

Effect of fluorodeoxyuridine on the sedimentation of nucleoids from HeLa cells in sucrose gradients.

PubMed

Synzynys, B I; Brozmanová, J; Saenko, A S

1987-01-01

Sedimentation properties of nucleoids from HeLa cells cultured for 6 or 24 h with 10(-6) M fluorodeoxyuridine (FdUrd) were studied in neutral sucrose gradients. Independently on the presence and concentrations of ethidium bromide in the gradient, nucleoids from FdUrd treated cells sedimented farther than those from untreated cells. However, the maximum relaxation of supercoiled DNA, observed at the concentration of 5 micrograms/ml of ethidium bromide, was significantly lower in cells treated with FdUrd, which indicated that prior incubation with FdUrd did not increase the degree of DNA supercoiling but altered by some way the conformation of DNA in nucleus. Previously we have found, that treatment of HeLa cells with FdUrd resulted in the stimulation of DNA synthesis, which proved to be resistant to ultraviolet and gamma-irradiation. From the present results it is possible to suggest, that alterations of chromatine structure should be included in facilitating of DNA synthesis on DNA template damaged by ultraviolet or gamma irradiation.

Analysis of Oligonucleotide DNA Binding and Sedimentation Properties of Montmorillonite Clay Using Ultraviolet Light Spectroscopy

PubMed Central

Beall, Gary W.; Sowersby, Drew S.; Roberts, Rachel D.; Robson, Michael H.; Lewis, L. Kevin

2009-01-01

Smectite clays such as montmorillonite form complexes with a variety of biomolecules, including the nucleic acids DNA and RNA. Most previous studies of DNA adsorption onto clay have relied upon spectrophotometric analysis after separation of free nucleic acids from bound complexes by centrifugation. In the current work we demonstrate that such studies produce a consistent error due to (a) incomplete sedimentation of montmorillonite and (b) strong absorbance of the remaining clay at 260 nm. Clay sedimentation efficiency was strongly dependent upon cation concentration (Na+ or Mg2+) and on the level of dispersion of the original suspension. An improved clay:DNA adsorption assay was developed and utilized to assess the impact of metal counterions on binding of single-stranded DNA to montmorillonite. X-ray diffraction demonstrated, for the first time, formation of intercalated structures consistent with orientation of the DNA strands parallel to the clay surface. Observed gallery spacings were found to closely match values calculated utilizing atomistic modeling techniques. PMID:19061334

Tailored protein encapsulation into a DNA host using geometrically organized supramolecular interactions

NASA Astrophysics Data System (ADS)

Sprengel, Andreas; Lill, Pascal; Stegemann, Pierre; Bravo-Rodriguez, Kenny; Schöneweiß, Elisa-C.; Merdanovic, Melisa; Gudnason, Daniel; Aznauryan, Mikayel; Gamrad, Lisa; Barcikowski, Stephan; Sanchez-Garcia, Elsa; Birkedal, Victoria; Gatsogiannis, Christos; Ehrmann, Michael; Saccà, Barbara

2017-02-01

The self-organizational properties of DNA have been used to realize synthetic hosts for protein encapsulation. However, current strategies of DNA-protein conjugation still limit true emulation of natural host-guest systems, whose formation relies on non-covalent bonds between geometrically matching interfaces. Here we report one of the largest DNA-protein complexes of semisynthetic origin held in place exclusively by spatially defined supramolecular interactions. Our approach is based on the decoration of the inner surface of a DNA origami hollow structure with multiple ligands converging to their corresponding binding sites on the protein surface with programmable symmetry and range-of-action. Our results demonstrate specific host-guest recognition in a 1:1 stoichiometry and selectivity for the guest whose size guarantees sufficient molecular diffusion preserving short intermolecular distances. DNA nanocontainers can be thus rationally designed to trap single guest molecules in their native form, mimicking natural strategies of molecular recognition and anticipating a new method of protein caging.

Chemical Modifications that Affect Nutritional and Functional Properties of Proteins.

ERIC Educational Resources Information Center

Richardson, T.; Kester, J. J.

1984-01-01

Discusses chemical alterations of selected amino acids resulting from environmental effects (photooxidations, pH extremes, thermally induced effects). Also dicusses use of intentional chemical derivatizations of various functional groups in amino acid residue side chains and how recombinant DNA techniques might be useful in structure/function…

Molecular and functional characterization of single-box high-mobility group B (HMGB) chromosomal protein from Aedes aegypti.

PubMed

de Abreu da Silva, Isabel Caetano; Vicentino, Amanda Roberta Revoredo; Dos Santos, Renata Coutinho; da Fonseca, Rodrigo Nunes; de Mendonça Amarante, Anderson; Carneiro, Vitor Coutinho; de Amorim Pinto, Marcia; Aguilera, Estefania Anahi; Mohana-Borges, Ronaldo; Bisch, Paulo Mascarello; da Silva-Neto, Mario Alberto Cardoso; Fantappié, Marcelo Rosado

2018-05-30

High-mobility group B (HMGB) proteins have highly conserved, unique DNA-binding domains, HMG boxes, that can bind non-B-type DNA structures, such as bent, kinked and unwound structures, with high affinity. HMGB proteins also promote DNA bending, looping and unwinding. In this study, we determined the role of the Aedes aegypti single HMG-box domain protein AaHMGB; characterized its structure, spatiotemporal expression levels, subcellular localization, and nucleic acid binding activities; and compared these properties with those of its double-HMG-box counterpart protein, AaHMGB1. Via qRT-PCR, we showed that AaHMGB is expressed at much higher levels than AaHMGB1 throughout mosquito development. In situ hybridization results suggested a role for AaHMGB and AaHMGB1 during embryogenesis. Immunolocalization in the midgut revealed that AaHMGB is exclusively nuclear. Circular dichroism and fluorescence spectroscopy analyses showed that AaHMGB exhibits common features of α-helical structures and is more stably folded than AaHMGB1, likely due to the presence of one or two HMG boxes. Using several DNA substrates or single-stranded RNAs as probes, we observed significant differences between AaHMGB and AaHMGB1 in terms of their binding patterns, activity and/or specificity. Importantly, we showed that the phosphorylation of AaHMGB plays a critical role in its DNA-binding activity. Our study provides additional insight into the roles of single- versus double-HMG-box-containing proteins in nucleic acid interactions for better understanding of mosquito development, physiology and homeostasis. Copyright © 2017. Published by Elsevier B.V.

Structural and biophysical properties of h-FANCI ARM repeat protein.

PubMed

Siddiqui, Mohd Quadir; Choudhary, Rajan Kumar; Thapa, Pankaj; Kulkarni, Neha; Rajpurohit, Yogendra S; Misra, Hari S; Gadewal, Nikhil; Kumar, Satish; Hasan, Syed K; Varma, Ashok K

2017-11-01

Fanconi anemia complementation groups - I (FANCI) protein facilitates DNA ICL (Inter-Cross-link) repair and plays a crucial role in genomic integrity. FANCI is a 1328 amino acids protein which contains armadillo (ARM) repeats and EDGE motif at the C-terminus. ARM repeats are functionally diverse and evolutionarily conserved domain that plays a pivotal role in protein-protein and protein-DNA interactions. Considering the importance of ARM repeats, we have explored comprehensive in silico and in vitro approach to examine folding pattern. Size exclusion chromatography, dynamic light scattering (DLS) and glutaraldehyde crosslinking studies suggest that FANCI ARM repeat exist as monomer as well as in oligomeric forms. Circular dichroism (CD) and fluorescence spectroscopy results demonstrate that protein has predominantly α- helices and well-folded tertiary structure. DNA binding was analysed using electrophoretic mobility shift assay by autoradiography. Temperature-dependent CD, Fluorescence spectroscopy and DLS studies concluded that protein unfolds and start forming oligomer from 30°C. The existence of stable portion within FANCI ARM repeat was examined using limited proteolysis and mass spectrometry. The normal mode analysis, molecular dynamics and principal component analysis demonstrated that helix-turn-helix (HTH) motif present in ARM repeat is highly dynamic and has anti-correlated motion. Furthermore, FANCI ARM repeat has HTH structural motif which binds to double-stranded DNA.

GEN1/Yen1 and the SLX4 complex: solutions to the problem of Holliday junction resolution

PubMed Central

Svendsen, Jennifer M.; Harper, J. Wade

2010-01-01

Chromosomal double-strand breaks (DSBs) are considered to be among the most deleterious DNA lesions found in eukaryotic cells due to their propensity to promote genome instability. DSBs occur as a result of exogenous or endogenous DNA damage, and also occur during meiotic recombination. DSBs are often repaired through a process called homologous recombination (HR), which employs the sister chromatid in mitotic cells or the homologous chromosome in meiotic cells, as a template for repair. HR frequently involves the formation and resolution of four-way DNA structures referred to as the Holliday junction (HJ). Despite extensive study, the machinery and mechanisms used to process these structures in eukaryotes have remained poorly understood. Recent work has identified XPG and UvrC/GIY domain-containing structure-specific endonucleases that can symmetrically cleave HJs in vitro in a manner that allows for religation without additional processing, properties that are reminiscent of the classical RuvC HJ resolvase in bacteria. Genetic studies reveal potential roles for these HJ resolvases in repair after DNA damage and during meiosis. The stage is now set for a more comprehensive understanding of the specific roles these enzymes play in the response of cells to DSBs, collapsed replication forks, telomere dysfunction, and meiotic recombination. PMID:20203129

Impact of low-frequency hotspot mutation R282Q on the structure of p53 DNA-binding domain as revealed by crystallography at 1.54 Å resolution

PubMed Central

Tu, Chao; Tan, Yu-Hong; Shaw, Gary; Zhou, Zheng; Bai, Yawen; Luo, Ray; Ji, Xinhua

2008-01-01

Tumor suppressor p53 is a sequence-specific DNA-binding protein and its central DNA-binding domain (DBD) harbors six hotspots (Arg175, Gly245, Arg248, Arg249, Arg273 and Arg282) for human cancers. Here, the crystal structure of a low-frequency hotspot mutant, p53DBD(R282Q), is reported at 1.54 Å resolution together with the results of molecular-dynamics simulations on the basis of the structure. In addition to eliminating a salt bridge, the R282Q mutation has a significant impact on the properties of two DNA-binding loops (L1 and L3). The L1 loop is flexible in the wild type, but it is not flexible in the mutant. The L3 loop of the wild type is not flexible, whereas it assumes two conformations in the mutant. Molecular-dynamics simulations indicated that both conformations of the L3 loop are accessible under biological conditions. It is predicted that the elimination of the salt bridge and the inversion of the flexibility of L1 and L3 are directly or indirectly responsible for deactivating the tumor suppressor p53. PMID:18453682

Synthesis and studies of polypeptide materials: Self-assembled block copolypeptide amphiphiles, DNA-condensing block copolypeptides and membrane-interactive random copolypeptides

NASA Astrophysics Data System (ADS)

Wyrsta, Michael Dmytro

A new class of transition metal initiators for the controlled polymerization of alpha-aminoacid-N-carboxyanhydrides (alpha-NCAs), has been developed by Deming et al. This discovery has allowed for the synthesis of well-defined "protein-like" polymers. Using this chemistry we have made distinct block/random copolypeptides for biomedical applications. Drug delivery, gene delivery, and antimicrobial polymers were the focus of our research efforts. The motivation for the synthesis and study of synthetic polypeptide based materials comes from proteins. Natural proteins are able to adopt a staggeringly large amount of uniquely well-defined folded structures. These structures account for the diversity in properties of proteins. As catalysts (enzymes) natural proteins perform some of the most difficult chemistry with ease and precision at ambient pressures and temperatures. They also exhibit incredible structural properties that directly result from formation of complex hierarchical assemblies. Self-assembling block copolymers were synthesized with various compositions and architectures. In general, di- and tri-block amphiphiles were studied for their self-assembling properties. Both spherical and tubular vesicles were found to assemble from di- and tri-block amphiphiles, respectively. In addition to self-assembly, pH responsiveness was engineered into these amphiphiles by the incorporation of basic residues (lysine) into the hydrophobic block. Another form of self-assembly studied was the condensation of DNA using cationic block copolymers. It was found that cationic block copolymers could condense DNA into compact, ordered, water-soluble aggregates on the nanoscale. These aggregates sufficiently protected DNA from nucleases and yet were susceptible to proteases. These studies form the basis of a gene delivery platform. The ease with which NCAs are polymerized renders them completely amenable to parallel synthetic methods. We have employed this technique to discover new antimicrobial polypeptides. The polymers studied were themselves the antimicrobial agent, not a self-assembled aggregate that contained antibiotics. It was found that powerful antibacterial polymers could be readily prepared with simple binary compositions. Antibacterial activity was sensitive to copolymer composition, bacterial cell-wall type, and insensitive to chain length (within reason).

Comparative thermal and thermodynamic study of DNA chemically modified with antitumor drug cisplatin and its inactive analog transplatin.

PubMed

Lando, Dmitri Y; Chang, Chun-Ling; Fridman, Alexander S; Grigoryan, Inessa E; Galyuk, Elena N; Hsueh, Ya-Wei; Hu, Chin-Kun

2014-08-01

Antitumor activity of cisplatin is exerted by covalent binding to DNA. For comparison, studies of cisplatin-DNA complexes often employ the very similar but inactive transplatin. In this work, thermal and thermodynamic properties of DNA complexes with these compounds were studied using differential scanning calorimetry (DSC) and computer modeling. DSC demonstrates that cisplatin decreases thermal stability (melting temperature, Tm) of long DNA, and transplatin increases it. At the same time, both compounds decrease the enthalpy and entropy of the helix-coil transition, and the impact of transplatin is much higher. From Pt/nucleotide molar ratio rb=0.001, both compounds destroy the fine structure of DSC profile and increase the temperature melting range (ΔT). For cisplatin and transplatin, the dependences δTm vs rb differ in sign, while δΔT vs rb are positive for both compounds. The change in the parameter δΔT vs rb demonstrates the GC specificity in the location of DNA distortions. Our experimental results and calculations show that 1) in contrast to [Pt(dien)Cl]Cl, monofunctional adducts formed by transplatin decrease the thermal stability of long DNA at [Na(+)]>30mM; 2) interstrand crosslinks of cisplatin and transplatin only slightly increase Tm; 3) the difference in thermal stability of DNA complexes with cisplatin vs DNA complexes with transplatin mainly arises from the different thermodynamic properties of their intrastrand crosslinks. This type of crosslink appears to be responsible for the antitumor activity of cisplatin. At any [Na(+)] from interval 10-210mM, cisplatin and transplatin intrastrand crosslinks give rise to destabilization and stabilization, respectively. Copyright © 2014 Elsevier Inc. All rights reserved.

Effects of cytotoxic cis- and trans-diammine monochlorido platinum(II) complexes on selenium-dependent redox enzymes and DNA.

PubMed

Lemmerhirt, Heidi; Behnisch, Steven; Bodtke, Anja; Lillig, Christopher H; Pazderova, Lucia; Kasparkova, Jana; Brabec, Viktor; Bednarski, Patrick J

2018-01-01

Here we present the preparation of 14 pairs of cis- and trans-diammine monochlorido platinum(II) complexes, coordinated to heterocycles (i.e., imidazole, 2-methylimidazole and pyrazole) and linked to various acylhydrazones, which were designed as potential inhibitors of the selenium-dependent enzymes glutathione peroxidase 1 (GPx-1) and thioredoxin reductase 1 (TrxR-1). However, no inhibition of bovine GPx-1 and only weak inhibition of murine TrxR-1 was observed in in vitro assays. Nonetheless, the cis configured diammine monochlorido Pt(II) complexes exhibited cytotoxic and apoptotic properties on various human cancer cell lines, whereas the trans configured complexes generally showed weaker potency with a few exceptions. On the other hand, the trans complexes were generally more likely to lack cross-resistance to cisplatin than the cis analogues. Platinum was found bound to the nuclear DNA of cancer cells treated with representative Pt complexes, suggesting that DNA might be a possible target. Thus, detailed in vitro binding experiments with DNA were conducted. Interactions of the compounds with calf thymus DNA were investigated, including Pt binding kinetics, circular dichroism (CD) spectral changes, changes in DNA melting temperatures, unwinding of supercoiled plasmids and ethidium bromide displacement in DNA. The CD results indicate that the most active cis configured pyrazole-derived complex causes unique structural changes in the DNA compared to the other complexes as well as to those caused by cisplatin, suggesting a denaturation of the DNA structure. This may be important for the antiproliferative activity of this compound in the cancer cells. Copyright © 2017. Published by Elsevier Inc.

Insights into Watson-Crick/Hoogsteen breathing dynamics and damage repair from the solution structure and dynamic ensemble of DNA duplexes containing m1A.

PubMed

Sathyamoorthy, Bharathwaj; Shi, Honglue; Zhou, Huiqing; Xue, Yi; Rangadurai, Atul; Merriman, Dawn K; Al-Hashimi, Hashim M

2017-05-19

In the canonical DNA double helix, Watson-Crick (WC) base pairs (bps) exist in dynamic equilibrium with sparsely populated (∼0.02-0.4%) and short-lived (lifetimes ∼0.2-2.5 ms) Hoogsteen (HG) bps. To gain insights into transient HG bps, we used solution-state nuclear magnetic resonance spectroscopy, including measurements of residual dipolar couplings and molecular dynamics simulations, to examine how a single HG bp trapped using the N1-methylated adenine (m1A) lesion affects the structural and dynamic properties of two duplexes. The solution structure and dynamic ensembles of the duplexes reveals that in both cases, m1A forms a m1A•T HG bp, which is accompanied by local and global structural and dynamic perturbations in the double helix. These include a bias toward the BI backbone conformation; sugar repuckering, major-groove directed kinking (∼9°); and local melting of neighboring WC bps. These results provide atomic insights into WC/HG breathing dynamics in unmodified DNA duplexes as well as identify structural and dynamic signatures that could play roles in m1A recognition and repair. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Insights into Watson–Crick/Hoogsteen breathing dynamics and damage repair from the solution structure and dynamic ensemble of DNA duplexes containing m1A

PubMed Central

Sathyamoorthy, Bharathwaj; Shi, Honglue; Zhou, Huiqing; Xue, Yi; Rangadurai, Atul; Merriman, Dawn K.

2017-01-01

Abstract In the canonical DNA double helix, Watson–Crick (WC) base pairs (bps) exist in dynamic equilibrium with sparsely populated (∼0.02–0.4%) and short-lived (lifetimes ∼0.2–2.5 ms) Hoogsteen (HG) bps. To gain insights into transient HG bps, we used solution-state nuclear magnetic resonance spectroscopy, including measurements of residual dipolar couplings and molecular dynamics simulations, to examine how a single HG bp trapped using the N1-methylated adenine (m1A) lesion affects the structural and dynamic properties of two duplexes. The solution structure and dynamic ensembles of the duplexes reveals that in both cases, m1A forms a m1A•T HG bp, which is accompanied by local and global structural and dynamic perturbations in the double helix. These include a bias toward the BI backbone conformation; sugar repuckering, major-groove directed kinking (∼9°); and local melting of neighboring WC bps. These results provide atomic insights into WC/HG breathing dynamics in unmodified DNA duplexes as well as identify structural and dynamic signatures that could play roles in m1A recognition and repair. PMID:28369571

The HIV-1 transcriptional activator Tat has potent nucleic acid chaperoning activities in vitro.

PubMed

Kuciak, Monika; Gabus, Caroline; Ivanyi-Nagy, Roland; Semrad, Katharina; Storchak, Roman; Chaloin, Olivier; Muller, Sylviane; Mély, Yves; Darlix, Jean-Luc

2008-06-01

The human immunodeficiency virus type 1 (HIV-1) is a primate lentivirus that causes the acquired immunodeficiency syndrome (AIDS). In addition to the virion structural proteins and enzyme precursors, that are Gag, Env and Pol, HIV-1 encodes several regulatory proteins, notably a small nuclear transcriptional activator named Tat. The Tat protein is absolutely required for virus replication since it controls proviral DNA transcription to generate the full-length viral mRNA. Tat can also regulate mRNA capping and splicing and was recently found to interfere with the cellular mi- and siRNA machinery. Because of its extensive interplay with nucleic acids, and its basic and disordered nature we speculated that Tat had nucleic acid-chaperoning properties. This prompted us to examine in vitro the nucleic acid-chaperoning activities of Tat and Tat peptides made by chemical synthesis. Here we report that Tat has potent nucleic acid-chaperoning activities according to the standard DNA annealing, DNA and RNA strand exchange, RNA ribozyme cleavage and trans-splicing assays. The active Tat(44-61) peptide identified here corresponds to the smallest known sequence with DNA/RNA chaperoning properties.

The HIV-1 transcriptional activator Tat has potent nucleic acid chaperoning activities in vitro

PubMed Central

Kuciak, Monika; Gabus, Caroline; Ivanyi-Nagy, Roland; Semrad, Katharina; Storchak, Roman; Chaloin, Olivier; Muller, Sylviane; Mély, Yves; Darlix, Jean-Luc

2008-01-01

The human immunodeficiency virus type 1 (HIV-1) is a primate lentivirus that causes the acquired immunodeficiency syndrome (AIDS). In addition to the virion structural proteins and enzyme precursors, that are Gag, Env and Pol, HIV-1 encodes several regulatory proteins, notably a small nuclear transcriptional activator named Tat. The Tat protein is absolutely required for virus replication since it controls proviral DNA transcription to generate the full-length viral mRNA. Tat can also regulate mRNA capping and splicing and was recently found to interfere with the cellular mi- and siRNA machinery. Because of its extensive interplay with nucleic acids, and its basic and disordered nature we speculated that Tat had nucleic acid-chaperoning properties. This prompted us to examine in vitro the nucleic acid-chaperoning activities of Tat and Tat peptides made by chemical synthesis. Here we report that Tat has potent nucleic acid-chaperoning activities according to the standard DNA annealing, DNA and RNA strand exchange, RNA ribozyme cleavage and trans-splicing assays. The active Tat(44–61) peptide identified here corresponds to the smallest known sequence with DNA/RNA chaperoning properties. PMID:18442994

Genome-wide survey of DNA-binding proteins in Arabidopsis thaliana: analysis of distribution and functions.

PubMed

Malhotra, Sony; Sowdhamini, Ramanathan

2013-08-01

The interaction of proteins with their respective DNA targets is known to control many high-fidelity cellular processes. Performing a comprehensive survey of the sequenced genomes for DNA-binding proteins (DBPs) will help in understanding their distribution and the associated functions in a particular genome. Availability of fully sequenced genome of Arabidopsis thaliana enables the review of distribution of DBPs in this model plant genome. We used profiles of both structure and sequence-based DNA-binding families, derived from PDB and PFam databases, to perform the survey. This resulted in 4471 proteins, identified as DNA-binding in Arabidopsis genome, which are distributed across 300 different PFam families. Apart from several plant-specific DNA-binding families, certain RING fingers and leucine zippers also had high representation. Our search protocol helped to assign DNA-binding property to several proteins that were previously marked as unknown, putative or hypothetical in function. The distribution of Arabidopsis genes having a role in plant DNA repair were particularly studied and noted for their functional mapping. The functions observed to be overrepresented in the plant genome harbour DNA-3-methyladenine glycosylase activity, alkylbase DNA N-glycosylase activity and DNA-(apurinic or apyrimidinic site) lyase activity, suggesting their role in specialized functions such as gene regulation and DNA repair.

Structural effects of TiO2 nanoparticles and doxorubicin on DNA and their antiproliferative roles in T47D and MCF7 cells.

PubMed

Hekmat, Azadeh; Saboury, Ali Akbar; Divsalar, Adeleh; Seyedarabi, Arefeh

2013-07-01

The structural changes in DNA caused by the combined effects of TiO2 nanoparticles (TiO2 NPs) and doxorubicin (DOX) were investigated along with their corresponding inhibitory roles in the growth of T47D and MCF7 cells. The UV-visible titration studies showed that DOX+ TiO2 NPs could form a novel complex with DNA. The data also reveal that the TiO2-DOX complex forms through a 1:4 stoichiometric ratio in solution. The values of binding constants reveal that DOX+TiO2 NPs interact more strongly with DNA as compared to TiO2 NPs or DOX alone. CD data show that DOX+TiO2 NPs can noticeably cause disturbance on DNA structure compared to TiO2 NPs or DOX alone, considering that DNA is relatively thermally stable in the condition used. The anticancer property of 0.3 µM DOX+ 60 µM TiO2 NPs and 0.4 µM DOX+ 670 µM TiO2 NPs by MTT assay and DAPI stain demonstrates that this combination can tremendously diminish proliferation of T47D and MCF7cells compared to DOX or TiO2 NPs alone. The UV-Vis absorption spectroscopy, flow cytometry and fluorescence microscopy experiments show much more enhancement of DOX uptake through the use of TiO2 NPs. These results reveal that DOX+TiO2 NPs could proffer a novel strategy for the development of promising and efficient chemotherapy agents.

mtDNA variation in caste populations of Andhra Pradesh, India.

PubMed

Bamshad, M; Fraley, A E; Crawford, M H; Cann, R L; Busi, B R; Naidu, J M; Jorde, L B

1996-02-01

Various anthropological analyses have documented extensive regional variation among populations on the subcontinent of India using morphological, protein, blood group, and nuclear DNA polymorphisms. These patterns are the product of complex population structure (genetic drift, gene flow) and a population history noted for numerous branching events. As a result, the interpretation of relationships among caste populations of South India and between Indians and continental populations remains controversial. The Hindu caste system is a general model of genetic differentiation among endogamous populations stratified by social forces (e.g., religion and occupation). The mitochondrial DNA (mtDNA) molecule has unique properties that facilitate the exploration of population structure. We analyzed 36 Hindu men born in Andhra Pradesh who were unrelated matrilineally through at least 3 generations and who represent 4 caste populations: Brahmin (9), Yadava (10), Kapu (7), and Relli (10). Individuals from Africa (36), Asia (36), and Europe (36) were sampled for comparison. A 200-base-pair segment of hypervariable segment 2 (HVS2) of the mtDNA control region was sequenced in all individuals. In the Indian castes 25 distinct haplotypes are identified. Aside from the Cambridge reference sequence, only two haplotypes are shared between caste populations. Middle castes form a highly supported cluster in a neighbor-joining network. Mean nucleotide diversity within each caste is 0.015, 0.012, 0.011, and 0.012 for the Brahmin, Yadava, Kapu, and Relli, respectively. mtDNA variation is highly structured between castes (GST = 0.17; p < 0.002). The effects of social structure on mtDNA variation are much greater than those on variation measured by traditional markers. Explanations for this discordance include (1) the higher resolving power of mtDNA, (2) sex-dependent gene flow, (3) differences in male and female effective population sizes, and (4) elements of the kinship structure. Thirty distinct haplotypes are found in Africans, 17 in Asians, and 13 in Europeans. Mean nucleotide diversity is 0.019, 0.014, 0.009, and 0.007 for Africans, Indians, Asians, and Europeans, respectively. These populations are highly structured geographically (GST = 0.15; p < 0.001). The caste populations of Andhra Pradesh cluster more often with Africans than with Asians or Europeans. This is suggestive of admixture with African populations.

Examining multi-component DNA-templated nanostructures as imaging agents

NASA Astrophysics Data System (ADS)

Jaganathan, Hamsa

2011-12-01

Magnetic resonance imaging (MRI) is the leading non-invasive tool for disease imaging and diagnosis. Although MRI exhibits high spatial resolution for anatomical features, the contrast resolution is low. Imaging agents serve as an aid to distinguish different types of tissues within images. Gadolinium chelates, which are considered first generation designs, can be toxic to health, while ultra-small, superparamagnetic nanoparticles (NPs) have low tissue-targeting efficiency and rapid bio-distribution, resulting to an inadequate detection of the MRI signal and enhancement of image contrast. In order to improve the utility of MRI agents, the challenge in composition and structure needs to be addressed. One-dimensional (1D), superparamagnetic nanostructures have been reported to enhance magnetic and in vivo properties and therefore has a potential to improve contrast enhancement in MRI images. In this dissertation, the structure of 1D, multi-component NP chains, scaffolded on DNA, were pre-clinically examined as potential MRI agents. First, research was focused on characterizing and understanding the mechanism of proton relaxation for DNA-templated NP chains using nuclear magnetic resonance (NMR) spectrometry. Proton relaxation and transverse relaxivity were higher in multi-component NP chains compared to disperse NPs, indicating the arrangement of NPs on a 1D structure improved proton relaxation sensitivity. Second, in vitro evaluation for potential issues in toxicity and contrast efficiency in tissue environments using a 3 Tesla clinical MRI scanner was performed. Cell uptake of DNA-templated NP chains was enhanced after encapsulating the nanostructure with layers of polyelectrolytes and targeting ligands. Compared to dispersed NPs, DNA-templated NP chains improved MRI contrast in both the epithelial basement membrane and colon cancer tumors scaffolds. The last part of the project was focused on developing a novel MRI agent that detects changes in DNA methylation levels. The findings from this dissertation suggest that the structural arrangement of NPs on DNA significantly influenced their function and utility as MRI agents.

Mechanisms of Insertion of dCTP and dTTP Opposite the DNA Lesion O6-Methyl-2'-deoxyguanosine by Human DNA Polymerase η.

PubMed

Patra, Amitraj; Zhang, Qianqian; Guengerich, F Peter; Egli, Martin

2016-11-11

O 6 -Methyl-2'-deoxyguanosine (O 6 -MeG) is a ubiquitous DNA lesion, formed not only by xenobiotic carcinogens but also by the endogenous methylating agent S-adenosylmethionine. It can introduce mutations during DNA replication, with different DNA polymerases displaying different ratios of correct or incorrect incorporation opposite this nucleoside. Of the "translesion" Y-family human DNA polymerases (hpols), hpol η is most efficient in incorporating equal numbers of correct and incorrect C and T bases. However, the mechanistic basis for this specific yet indiscriminate activity is not known. To explore this question, we report biochemical and structural analysis of the catalytic core of hpol η. Activity assays showed the truncated form displayed similar misincorporation properties as the full-length enzyme, incorporating C and T equally and extending from both. X-ray crystal structures of both dC and dT paired with O 6 -MeG were solved in both insertion and extension modes. The structures revealed a Watson-Crick-like pairing between O 6 -MeG and 2"-deoxythymidine-5"-[(α, β)-imido]triphosphate (approximating dT) at both the insertion and extension stages with formation of two H-bonds. Conversely, both the structures with O 6 - MeG opposite dCTP and dC display sheared configuration of base pairs but to different degrees, with formation of two bifurcated H-bonds and two single H-bonds in the structures trapped in the insertion and extension states, respectively. The structural data are consistent with the observed tendency of hpol η to insert both dC and dT opposite the O 6 -MeG lesion with similar efficiencies. Comparison of the hpol η active site configurations with either O 6 -MeG:dC or O 6 -MeG:dT bound compared with the corresponding situations in structures of complexes of Sulfolobus solfataricus Dpo4, a bypass pol that favors C relative to T by a factor of ∼4, helps rationalize the more error-prone synthesis opposite the lesion by hpol η. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Mechanisms of Insertion of dCTP and dTTP Opposite the DNA Lesion O6-Methyl-2′-deoxyguanosine by Human DNA Polymerase η*

PubMed Central

Patra, Amitraj; Zhang, Qianqian; Guengerich, F. Peter; Egli, Martin

2016-01-01

O6-Methyl-2′-deoxyguanosine (O6-MeG) is a ubiquitous DNA lesion, formed not only by xenobiotic carcinogens but also by the endogenous methylating agent S-adenosylmethionine. It can introduce mutations during DNA replication, with different DNA polymerases displaying different ratios of correct or incorrect incorporation opposite this nucleoside. Of the “translesion” Y-family human DNA polymerases (hpols), hpol η is most efficient in incorporating equal numbers of correct and incorrect C and T bases. However, the mechanistic basis for this specific yet indiscriminate activity is not known. To explore this question, we report biochemical and structural analysis of the catalytic core of hpol η. Activity assays showed the truncated form displayed similar misincorporation properties as the full-length enzyme, incorporating C and T equally and extending from both. X-ray crystal structures of both dC and dT paired with O6-MeG were solved in both insertion and extension modes. The structures revealed a Watson-Crick-like pairing between O6-MeG and 2"-deoxythymidine-5"-[(α, β)-imido]triphosphate (approximating dT) at both the insertion and extension stages with formation of two H-bonds. Conversely, both the structures with O6- MeG opposite dCTP and dC display sheared configuration of base pairs but to different degrees, with formation of two bifurcated H-bonds and two single H-bonds in the structures trapped in the insertion and extension states, respectively. The structural data are consistent with the observed tendency of hpol η to insert both dC and dT opposite the O6-MeG lesion with similar efficiencies. Comparison of the hpol η active site configurations with either O6-MeG:dC or O6-MeG:dT bound compared with the corresponding situations in structures of complexes of Sulfolobus solfataricus Dpo4, a bypass pol that favors C relative to T by a factor of ∼4, helps rationalize the more error-prone synthesis opposite the lesion by hpol η. PMID:27694439

Mechanical properties and cell-culture characteristics of a polycaprolactone kagome-structure scaffold fabricated by a precision extruding deposition system.

PubMed

Lee, Se-Hwan; Cho, Yong Sang; Hong, Myoung Wha; Lee, Bu-Kyu; Park, Yongdoo; Park, Sang-Hyug; Kim, Young Yul; Cho, Young-Sam

2017-09-13

To enhance the mechanical properties of three-dimensional (3D) scaffolds used for bone regeneration in tissue engineering, many researchers have studied their structure and chemistry. In the structural engineering field, the kagome structure has been known to have an excellent relative strength. In this study, to enhance the mechanical properties of a synthetic polymer scaffold used for tissue engineering, we applied the 3D kagome structure to a porous scaffold for bone regeneration. Prior to fabricating the biocompatible-polymer scaffold, the ideal kagome structure, which was manufactured by a 3D printer of the digital light processing type, was compared with a grid-structure, which was used as the control group, using a compressive experiment. A polycaprolactone (PCL) kagome-structure scaffold was successfully fabricated by additive manufacturing using a 3D printer with a precision extruding deposition head. To assess the physical characteristics of the fabricated PCL-kagome-structure scaffold, we analyzed its porosity, pore size, morphological structure, surface roughness, compressive stiffness, and mechanical bending properties. The results showed that, the mechanical properties of proposed kagome-structure scaffold were superior to those of a grid-structure scaffold. Moreover, Sarcoma osteogenic (Saos-2) cells were used to evaluate the characteristics of in vitro cell proliferation. We carried out cell counting kit-8 (CCK-8) and DNA contents assays. Consequently, the cell proliferation of the kagome-structure scaffold was increased; this could be because the surface roughness of the kagome-structure scaffold enhances initial cell attachment.

The development of bis(hydroxymethyl)pyrrole analogs as bifunctional DNA cross-linking agents and their chemotherapeutic potential.

PubMed

Su, Tsann-Long; Lee, Te-Chang; Kakadiya, Rajesh

2013-11-01

Bifunctional DNA cross-linking agents are widely used as chemotherapeutic agents in clinics. The advance in the development of these agents as potential antitumor agents has generated various types of bis(hydroxymethyl)pyrrole analogs. In order to develop highly effective anticancer agents, it is necessary to understand the chemophysical properties, structure-activity relationships, therapeutic potency, toxicity/safety, and pharmacokinetics of these DNA cross-linking agents. This review presents an overview of the recent advances in developing various types of bis(hydroxymethyl)pyrrole analogs with potential antitumor activity to provide more information for future drug design and strategies for combination chemotherapy. The rational drug design, chemical syntheses, antitumor activity, mechanism of action, and development of combined chemotherapy regimens, including a DNA repair inhibitor, are discussed. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

Identification of a Herbal Powder by Deoxyribonucleic Acid Barcoding and Structural Analyses.

PubMed

Sheth, Bhavisha P; Thaker, Vrinda S

2015-10-01

Authentic identification of plants is essential for exploiting their medicinal properties as well as to stop the adulteration and malpractices with the trade of the same. To identify a herbal powder obtained from a herbalist in the local vicinity of Rajkot, Gujarat, using deoxyribonucleic acid (DNA) barcoding and molecular tools. The DNA was extracted from a herbal powder and selected Cassia species, followed by the polymerase chain reaction (PCR) and sequencing of the rbcL barcode locus. Thereafter the sequences were subjected to National Center for Biotechnology Information (NCBI) basic local alignment search tool (BLAST) analysis, followed by the protein three-dimension structure determination of the rbcL protein from the herbal powder and Cassia species namely Cassia fistula, Cassia tora and Cassia javanica (sequences obtained in the present study), Cassia Roxburghii, and Cassia abbreviata (sequences retrieved from Genbank). Further, the multiple and pairwise structural alignment were carried out in order to identify the herbal powder. The nucleotide sequences obtained from the selected species of Cassia were submitted to Genbank (Accession No. JX141397, JX141405, JX141420). The NCBI BLAST analysis of the rbcL protein from the herbal powder showed an equal sequence similarity (with reference to different parameters like E value, maximum identity, total score, query coverage) to C. javanica and C. roxburghii. In order to solve the ambiguities of the BLAST result, a protein structural approach was implemented. The protein homology models obtained in the present study were submitted to the protein model database (PM0079748-PM0079753). The pairwise structural alignment of the herbal powder (as template) and C. javanica and C. roxburghii (as targets individually) revealed a close similarity of the herbal powder with C. javanica. A strategy as used here, incorporating the integrated use of DNA barcoding and protein structural analyses could be adopted, as a novel rapid and economic procedure, especially in cases when protein coding loci are considered. Authentic identification of plants is essential for exploiting their medicinal properties as well as to stop the adulteration and malpractices with the trade of the same. A herbal powder was obtained from a herbalist in the local vicinity of Rajkot, Gujarat. An integrated approach using DNA barcoding and structural analyses was carried out to identify the herbal powder. The herbal powder was identified as Cassia javanica L.

Evaluation of Methods for the Extraction of DNA from Drinking Water Distribution System Biofilms

PubMed Central

Hwang, Chiachi; Ling, Fangqiong; Andersen, Gary L.; LeChevallier, Mark W.; Liu, Wen-Tso

2012-01-01

While drinking water biofilms have been characterized in various drinking water distribution systems (DWDS), little is known about the impact of different DNA extraction methods on the subsequent analysis of microbial communities in drinking water biofilms. Since different DNA extraction methods have been shown to affect the outcome of microbial community analysis in other environments, it is necessary to select a DNA extraction method prior to the application of molecular tools to characterize the complex microbial ecology of the DWDS. This study compared the quantity and quality of DNA yields from selected DWDS bacteria with different cell wall properties using five widely used DNA extraction methods. These were further selected and evaluated for their efficiency and reproducibility of DNA extraction from DWDS samples. Terminal restriction fragment length analysis and the 454 pyrosequencing technique were used to interpret the differences in microbial community structure and composition, respectively, from extracted DNA. Such assessments serve as a concrete step towards the determination of an optimal DNA extraction method for drinking water biofilms, which can then provide a reliable comparison of the meta-analysis results obtained in different laboratories. PMID:22075624

The Composition and Structure of Biofilms Developed by Propionibacterium acnes Isolated from Cardiac Pacemaker Devices.

PubMed

Okuda, Ken-Ichi; Nagahori, Ryuichi; Yamada, Satomi; Sugimoto, Shinya; Sato, Chikara; Sato, Mari; Iwase, Tadayuki; Hashimoto, Kazuhiro; Mizunoe, Yoshimitsu

2018-01-01

The present study aimed to understand the biofilm formation mechanism of Propionibacterium acnes by analyzing the components and structure of the biofilms. P. acnes strains were isolated from the surface of explanted cardiac pacemaker devices that exhibited no clinical signs of infection. Culture tests using a simple stamp culture method (pressing pacemakers against the surface of agar plates) revealed frequent P. acnes colonization on the surface of cardiac pacemaker devices. P . acnes was isolated from 7/31 devices, and the isolates were categorized by multilocus sequence typing into five different sequence types (STs): ST4 (JK18.2), ST53 (JK17.1), ST69 (JK12.2 and JK13.1), ST124 (JK5.3), ST125 (JK6.2), and unknown ST (JK19.3). An in vitro biofilm formation assay using microtiter plates demonstrated that 5/7 isolates formed biofilms. Inhibitory effects of DNase I and proteinase K on biofilm formation varied among isolates. In contrast, dispersin B showed no inhibitory activity against all isolates. Three-dimensional live/dead imaging of P. acnes biofilms with different biochemical properties using confocal laser microscopy demonstrated different distributions and proportions of living and dead cells. Additionally, it was suggested that extracellular DNA (eDNA) plays a role in the formation of biofilms containing living cells. Ultrastructural analysis of P. acnes biofilms using a transmission electron microscope and atmospheric scanning electron microscope revealed leakage of cytoplasmic components along with cell lysis and fibrous structures of eDNA connecting cells. In conclusion, the biochemical properties and structures of the biofilms differed among P. acnes isolates. These findings may provide clues for establishing countermeasures against biofilm-associated infection by P. acnes .

The Composition and Structure of Biofilms Developed by Propionibacterium acnes Isolated from Cardiac Pacemaker Devices

PubMed Central

Okuda, Ken-ichi; Nagahori, Ryuichi; Yamada, Satomi; Sugimoto, Shinya; Sato, Chikara; Sato, Mari; Iwase, Tadayuki; Hashimoto, Kazuhiro; Mizunoe, Yoshimitsu

2018-01-01

The present study aimed to understand the biofilm formation mechanism of Propionibacterium acnes by analyzing the components and structure of the biofilms. P. acnes strains were isolated from the surface of explanted cardiac pacemaker devices that exhibited no clinical signs of infection. Culture tests using a simple stamp culture method (pressing pacemakers against the surface of agar plates) revealed frequent P. acnes colonization on the surface of cardiac pacemaker devices. P. acnes was isolated from 7/31 devices, and the isolates were categorized by multilocus sequence typing into five different sequence types (STs): ST4 (JK18.2), ST53 (JK17.1), ST69 (JK12.2 and JK13.1), ST124 (JK5.3), ST125 (JK6.2), and unknown ST (JK19.3). An in vitro biofilm formation assay using microtiter plates demonstrated that 5/7 isolates formed biofilms. Inhibitory effects of DNase I and proteinase K on biofilm formation varied among isolates. In contrast, dispersin B showed no inhibitory activity against all isolates. Three-dimensional live/dead imaging of P. acnes biofilms with different biochemical properties using confocal laser microscopy demonstrated different distributions and proportions of living and dead cells. Additionally, it was suggested that extracellular DNA (eDNA) plays a role in the formation of biofilms containing living cells. Ultrastructural analysis of P. acnes biofilms using a transmission electron microscope and atmospheric scanning electron microscope revealed leakage of cytoplasmic components along with cell lysis and fibrous structures of eDNA connecting cells. In conclusion, the biochemical properties and structures of the biofilms differed among P. acnes isolates. These findings may provide clues for establishing countermeasures against biofilm-associated infection by P. acnes. PMID:29491850

Probing the electrostatics and pharmacologic modulation of sequence-specific binding by the DNA-binding domain of the ETS-family transcription factor PU.1: a binding affinity and kinetics investigation

PubMed Central

Munde, Manoj; Poon, Gregory M. K.; Wilson, W. David

2013-01-01

Members of the ETS family of transcription factors regulate a functionally diverse array of genes. All ETS proteins share a structurally-conserved but sequence-divergent DNA-binding domain, known as the ETS domain. Although the structure and thermodynamics of the ETS-DNA complexes are well known, little is known about the kinetics of sequence recognition, a facet that offers potential insight into its molecular mechanism. We have characterized DNA binding by the ETS domain of PU.1 by biosensor-surface plasmon resonance (SPR). SPR analysis revealed a striking kinetic profile for DNA binding by the PU.1 ETS domain. At low salt concentrations, it binds high-affinity cognate DNA with a very slow association rate constant (≤105 M−1 s−1), compensated by a correspondingly small dissociation rate constant. The kinetics are strongly salt-dependent but mutually balance to produce a relatively weak dependence in the equilibrium constant. This profile contrasts sharply with reported data for other ETS domains (e.g., Ets-1, TEL) for which high-affinity binding is driven by rapid association (>107 M−1 s−1). We interpret this difference in terms of the hydration properties of ETS-DNA binding and propose that at least two mechanisms of sequence recognition are employed by this family of DNA-binding domain. Additionally, we use SPR to demonstrate the potential for pharmacological inhibition of sequence-specific ETS-DNA binding, using the minor groove-binding distamycin as a model compound. Our work establishes SPR as a valuable technique for extending our understanding of the molecular mechanisms of ETS-DNA interactions as well as developing potential small-molecule agents for biotechnological and therapeutic purposes. PMID:23416556

Self-assembly of polyelectrolyte surfactant complexes using large scale MD simulation

NASA Astrophysics Data System (ADS)

Goswami, Monojoy; Sumpter, Bobby

2014-03-01

Polyelectrolytes (PE) and surfactants are known to form interesting structures with varied properties in aqueous solutions. The morphological details of the PE-surfactant complexes depend on a combination of polymer backbone, electrostatic interactions and hydrophobic interactions. We study the self-assembly of cationic PE and anionic surfactants complexes in dilute condition. The importance of such complexes of PE with oppositely charged surfactants can be found in biological systems, such as immobilization of enzymes in polyelectrolyte complexes or nonspecific association of DNA with protein. Many useful properties of PE surfactant complexes come from the highly ordered structures of surfactant self-assembly inside the PE aggregate which has applications in industry. We do large scale molecular dynamics simulation using LAMMPS to understand the structure and dynamics of PE-surfactant systems. Our investigation shows highly ordered pearl-necklace structures that have been observed experimentally in biological systems. We investigate many different properties of PE-surfactant complexation for different parameter ranges that are useful for pharmaceutical, engineering and biological applications.

Structure, Stiffness and Substates of the Dickerson-Drew Dodecamer

PubMed Central

Dršata, Tomáš; Pérez, Alberto; Orozco, Modesto; Morozov, Alexandre V.; Šponer, Jiřĺ; Lankaš, Filip

2013-01-01

The Dickerson–Drew dodecamer (DD) d-[CGCGAATTCGCG]2 is a prototypic B-DNA molecule whose sequence-specific structure and dynamics have been investigated by many experimental and computational studies. Here, we present an analysis of DD properties based on extensive atomistic molecular dynamics (MD) simulations using different ionic conditions and water models. The 0.6–2.4-µs-long MD trajectories are compared to modern crystallographic and NMR data. In the simulations, the duplex ends can adopt an alternative base-pairing, which influences the oligomer structure. A clear relationship between the BI/BII backbone substates and the basepair step conformation has been identified, extending previous findings and exposing an interesting structural polymorphism in the helix. For a given end pairing, distributions of the basepair step coordinates can be decomposed into Gaussian-like components associated with the BI/BII backbone states. The nonlocal stiffness matrices for a rigid-base mechanical model of DD are reported for the first time, suggesting salient stiffness features of the central A-tract. The Riemann distance and Kullback–Leibler divergence are used for stiffness matrix comparison. The basic structural parameters converge very well within 300 ns, convergence of the BI/BII populations and stiffness matrices is less sharp. Our work presents new findings about the DD structural dynamics, mechanical properties, and the coupling between basepair and backbone configurations, including their statistical reliability. The results may also be useful for optimizing future force fields for DNA. PMID:23976886

Structure and specificity of the RNA-guided endonuclease Cas9 during DNA interrogation, target binding and cleavage

PubMed Central

Josephs, Eric A.; Kocak, D. Dewran; Fitzgibbon, Christopher J.; McMenemy, Joshua; Gersbach, Charles A.; Marszalek, Piotr E.

2015-01-01

CRISPR-associated endonuclease Cas9 cuts DNA at variable target sites designated by a Cas9-bound RNA molecule. Cas9's ability to be directed by single ‘guide RNA’ molecules to target nearly any sequence has been recently exploited for a number of emerging biological and medical applications. Therefore, understanding the nature of Cas9's off-target activity is of paramount importance for its practical use. Using atomic force microscopy (AFM), we directly resolve individual Cas9 and nuclease-inactive dCas9 proteins as they bind along engineered DNA substrates. High-resolution imaging allows us to determine their relative propensities to bind with different guide RNA variants to targeted or off-target sequences. Mapping the structural properties of Cas9 and dCas9 to their respective binding sites reveals a progressive conformational transformation at DNA sites with increasing sequence similarity to its target. With kinetic Monte Carlo (KMC) simulations, these results provide evidence of a ‘conformational gating’ mechanism driven by the interactions between the guide RNA and the 14th–17th nucleotide region of the targeted DNA, the stabilities of which we find correlate significantly with reported off-target cleavage rates. KMC simulations also reveal potential methodologies to engineer guide RNA sequences with improved specificity by considering the invasion of guide RNAs into targeted DNA duplex. PMID:26384421

One small step for Mot1; one giant leap for other Swi2/Snf2 enzymes?

PubMed Central

Viswanathan, Ramya; Auble, David T.

2011-01-01

The TATA-binding protein (TBP) is a major target for transcriptional regulation. Mot1, a Swi2/Snf2-related ATPase, dissociates TBP from DNA in an ATP dependent process. The experimental advantages of this relatively simple reaction have been exploited to learn more about how Swi2/Snf2 ATPases function biochemically. However, many unanswered questions remain and fundamental aspects of the Mot1 mechanism are still under debate. Here, we review the available data and integrate the results with structural and biochemical studies of related enzymes to derive a model for Mot1’s catalytic action consistent with the broad literature on enzymes in this family. We propose that the Mot1 ATPase domain is tethered to TBP by a flexible, spring-like linker of alpha helical hairpins. The linker juxtaposes the ATPase domain such that it can engage duplex DNA on one side of the TBP-DNA complex. This allows the ATPase to employ short-range, nonprocessive ATP-driven DNA tracking to pull or push TBP off its DNA site. DNA translocation is a conserved property of ATPases in the broader enzyme family. As such, the model explains how a structurally and functionally conserved ATPase domain has been put to use in a very different context than other enzymes in the Swi2/Snf2 family. PMID:21658482

2011 Rita Schaffer lecture: nanoparticles for intracellular nucleic acid delivery.

PubMed

Green, Jordan J

2012-07-01

Nanoparticles are a promising technology for delivery of new types of therapeutics. A polymer library approach has allowed engineering of polymeric particles that are particularly effective for the delivery of DNA and siRNA to human cells. Certain chemical structural motifs, degradable linkages, hydrophobicity, and biophysical properties are key for successful intracellular delivery. Small differences to biomaterial structure, and especially the type of degradable linkage in the polymers, can be critical for successful delivery of siRNA vs. DNA. Furthermore, subtle changes to biomaterial structure can facilitate cell-type gene delivery specificity between human brain cancer cells and healthy cells as well as between human retinal endothelial cells and epithelial cells. These polymeric nanoparticles are effective for nucleic acid delivery in a broad range of human cell types and have applications to regenerative medicine, ophthalmology, and cancer among many other biomedical research areas.

Ru(II)-polypyridyl surface functionalised gold nanoparticles as DNA targeting supramolecular structures and luminescent cellular imaging agents.

PubMed

Martínez-Calvo, Miguel; Orange, Kim N; Elmes, Robert B P; la Cour Poulsen, Bjørn; Williams, D Clive; Gunnlaugsson, Thorfinnur

2016-01-07

The development of Ru(II) functionalized gold nanoparticles 1–3·AuNP is described. These systems were found to be mono-disperse with a hydrodynamic radius of ca. 15 nm in water but gave rise to the formation of higher order structures in buffered solution. The interaction of 1–3·AuNP with DNA was also studied by spectroscopic and microscopic methods and suggested the formation of large self-assembly structures in solution. The uptake of 1–3·AuNP by cancer cells was studied using both confocal fluorescence as well as transmission electron microscopy (TEM), with the aim of investigating their potential as tools for cellular biology. These systems displaying a non-toxic profile with favourable photophysical properties may have application across various biological fields including diagnostics and therapeutics.

Molecular structures guide the engineering of chromatin

PubMed Central

Tekel, Stefan J.

2017-01-01

Abstract Chromatin is a system of proteins, RNA, and DNA that interact with each other to organize and regulate genetic information within eukaryotic nuclei. Chromatin proteins carry out essential functions: packing DNA during cell division, partitioning DNA into sub-regions within the nucleus, and controlling levels of gene expression. There is a growing interest in manipulating chromatin dynamics for applications in medicine and agriculture. Progress in this area requires the identification of design rules for the chromatin system. Here, we focus on the relationship between the physical structure and function of chromatin proteins. We discuss key research that has elucidated the intrinsic properties of chromatin proteins and how this information informs design rules for synthetic systems. Recent work demonstrates that chromatin-derived peptide motifs are portable and in some cases can be customized to alter their function. Finally, we present a workflow for fusion protein design and discuss best practices for engineering chromatin to assist scientists in advancing the field of synthetic epigenetics. PMID:28609787

Mononuclear zinc(II) complexes of 2-((2-(piperazin-1-yl)ethylimino)methyl)-4-substituted phenols: Synthesis, structural characterization, DNA binding and cheminuclease activities

NASA Astrophysics Data System (ADS)

Ravichandran, J.; Gurumoorthy, P.; Karthick, C.; Kalilur Rahiman, A.

2014-03-01

Four new zinc(II) complexes [Zn(HL1-4)Cl2] (1-4), where HL1-4 = 2-((2-(piperazin-1-yl)ethylimino)methyl)-4-substituted phenols, have been isolated and fully characterized using various spectro-analytical techniques. The X-ray crystal structure of complex 4 shows the distorted trigonal-bipyramidal coordination geometry around zinc(II) ion. The crystal packing is stabilized by intermolecular NH⋯O hydrogen bonding interaction. The complexes display no d-d electronic band in the visible region due to d10 electronic configuration of zinc(II) ion. The electrochemical properties of the synthesized ligands and their complexes exhibit similar voltammogram at reduction potential due to electrochemically innocent Zn(II) ion, which evidenced that the electron transfer is due to the nature of the ligand. Binding interaction of complexes with calf thymus DNA was studied by UV-Vis absorption titration, viscometric titration and cyclic voltammetry. All complexes bind with CT DNA by intercalation, giving the binding affinity in the order of 2 > 1 ≫ 3 > 4. The prominent cheminuclease activity of complexes on plasmid DNA (pBR322 DNA) was observed in the absence and presence of H2O2. Oxidative pathway reveals that the underlying mechanism involves hydroxyl radical.

DNA-binding proteins from marine bacteria expand the known sequence diversity of TALE-like repeats

PubMed Central

de Lange, Orlando; Wolf, Christina; Thiel, Philipp; Krüger, Jens; Kleusch, Christian; Kohlbacher, Oliver; Lahaye, Thomas

2015-01-01

Transcription Activator-Like Effectors (TALEs) of Xanthomonas bacteria are programmable DNA binding proteins with unprecedented target specificity. Comparative studies into TALE repeat structure and function are hindered by the limited sequence variation among TALE repeats. More sequence-diverse TALE-like proteins are known from Ralstonia solanacearum (RipTALs) and Burkholderia rhizoxinica (Bats), but RipTAL and Bat repeats are conserved with those of TALEs around the DNA-binding residue. We study two novel marine-organism TALE-like proteins (MOrTL1 and MOrTL2), the first to date of non-terrestrial origin. We have assessed their DNA-binding properties and modelled repeat structures. We found that repeats from these proteins mediate sequence specific DNA binding conforming to the TALE code, despite low sequence similarity to TALE repeats, and with novel residues around the BSR. However, MOrTL1 repeats show greater sequence discriminating power than MOrTL2 repeats. Sequence alignments show that there are only three residues conserved between repeats of all TALE-like proteins including the two new additions. This conserved motif could prove useful as an identifier for future TALE-likes. Additionally, comparing MOrTL repeats with those of other TALE-likes suggests a common evolutionary origin for the TALEs, RipTALs and Bats. PMID:26481363

A DNA network as an information processing system.

PubMed

Santini, Cristina Costa; Bath, Jonathan; Turberfield, Andrew J; Tyrrell, Andy M

2012-01-01

Biomolecular systems that can process information are sought for computational applications, because of their potential for parallelism and miniaturization and because their biocompatibility also makes them suitable for future biomedical applications. DNA has been used to design machines, motors, finite automata, logic gates, reaction networks and logic programs, amongst many other structures and dynamic behaviours. Here we design and program a synthetic DNA network to implement computational paradigms abstracted from cellular regulatory networks. These show information processing properties that are desirable in artificial, engineered molecular systems, including robustness of the output in relation to different sources of variation. We show the results of numerical simulations of the dynamic behaviour of the network and preliminary experimental analysis of its main components.

Multispectroscopic investigation of the interaction of BSA and DNA with the anticancer drug, N-(6-ferrocenyl-2-naphthoyl)-gamma-amino butyric acid methyl ester

NASA Astrophysics Data System (ADS)

Rajina, S. R.; Sudhi, Geethu; Austin, P.; Praveen, S. G.; Xavier, T. S.; Kenny, Peter T. M.; Binoy, J.

2018-05-01

The interaction of a drug with DNA and BSA play a great role in studying anti cancer activity and drug transport properties, which can be effectively, investigated using vibrational spectroscopy, UV visible spectroscopy and Fluorescence spectroscopy. The present work reports the structural features of N-(6-ferrocenyl-2-naphthoyl)-gamma-amino butyric acid Methyl ester (FNGABME) based on FTIR and FTRaman spectroscopy. The absorption and fluorescence spectroscopic methods were used to study the efficiency of the interaction of the compound FNGABME with BSA and DNA and also molecular docking were performed computationally to validate the results which shows that the title compound may exhibit inhibitory activity against the cancer cells.

Reconstruction of Toll-like receptor 9-mediated responses in HEK-Blue hTLR9 cells by transfection of human macrophage scavenger receptor 1 gene.

PubMed

Ohtsuki, Shozo; Takahashi, Yuki; Inoue, Takao; Takakura, Yoshinobu; Nishikawa, Makiya

2017-10-20

We used human Toll-like receptor 9 (hTLR9)-expressing HEK-Blue hTLR9 cells, which release secreted embryonic alkaline phosphatase (SEAP) upon response to CpG DNA, to evaluate the immunological properties of nucleic acid drug candidates. Our preliminary studies showed that phosphodiester CpG DNA hardly induced any SEAP secretion in HEK-Blue hTLR9 cells. In the current study, therefore, we developed HEK-Blue hTLR9 cells transduced with human macrophage scavenger receptor-1 (hMSR1), a cell-surface DNA receptor, and determined whether HEK-Blue hTLR9/hMSR1 cells respond to phosphorothioate (PS) CpG DNA and phosphodiester (PO) CpG DNA. We selected PS CpG2006, a single-stranded PO CpG DNA (ssCpG), and a tetrapod-like structured DNA (tetrapodna) containing ssCpG (tetraCpG) as model TLR9 ligands. Alexa Fluor 488-labeled ligands were used for flow cytometry. Unlike the mock-transfected HEK-Blue hTLR9 cells, the HEK-Blue hTLR9/hMSR1 cells efficiently took up all three CpG DNAs. SEAP release was almost proportional to the uptake. Treatment of HEK-Blue hTLR9/hMSR1 cells with an anti-hMSR1 antibody significantly reduced the uptake of ssCpG and tetraCpG. Collectively, reconstruction of TLR9-mediated responses to CpG DNA in HEK-Blue hTLR9 cells can be used to evaluate the toxicity of nucleic acid drug candidates with diverse physicochemical properties.

The Elusive Nature of Adaptive Mitochondrial DNA Evolution of an Arctic Lineage Prone to Frequent Introgression

PubMed Central

Melo-Ferreira, José; Vilela, Joana; Fonseca, Miguel M.; da Fonseca, Rute R.; Boursot, Pierre; Alves, Paulo C.

2014-01-01

Mitochondria play a fundamental role in cellular metabolism, being responsible for most of the energy production of the cell in the oxidative phosphorylation (OXPHOS) pathway. Mitochondrial DNA (mtDNA) encodes for key components of this process, but its direct role in adaptation remains far from understood. Hares (Lepus spp.) are privileged models to study the impact of natural selection on mitogenomic evolution because 1) species are adapted to contrasting environments, including arctic, with different metabolic pressures, and 2) mtDNA introgression from arctic into temperate species is widespread. Here, we analyzed the sequences of 11 complete mitogenomes (ten newly obtained) of hares of temperate and arctic origins (including two of arctic origin introgressed into temperate species). The analysis of patterns of codon substitutions along the reconstructed phylogeny showed evidence for positive selection in several codons in genes of the OXPHOS complexes, most notably affecting the arctic lineage. However, using theoretical models, no predictable effect of these differences was found on the structure and physicochemical properties of the encoded proteins, suggesting that the focus of selection may lie on complex interactions with nuclear encoded peptides. Also, a cloverleaf structure was detected in the control region only from the arctic mtDNA lineage, which may influence mtDNA replication and transcription. These results suggest that adaptation impacted the evolution of hare mtDNA and may have influenced the occurrence and consequences of the many reported cases of massive mtDNA introgression. However, the origin of adaptation remains elusive. PMID:24696399

Spectroscopic studies of the interaction of aspirin and its important metabolite, salicylate ion, with DNA, A·T and G·C rich sequences

NASA Astrophysics Data System (ADS)

Bathaie, S. Z.; Nikfarjam, L.; Rahmanpour, R.; Moosavi-Movahedi, A. A.

2010-12-01

Among different biological effects of acetylsalicylic acid (ASA), its anticancer property is controversial. Since ASA hydrolyzes rapidly to salicylic acid (SA), especially in the blood, interaction of both ASA and SA (as the small molecules) with ctDNA, oligo(dA·dT) 15 and oligo(dG·dC) 15, as a possible mechanism of their action, is investigated here. The results show that the rate of ASA hydrolysis in the absence and presence of ctDNA is similar. The spectrophotometric results indicate that both ASA and SA cooperatively bind to ctDNA. The binding constants ( K) are (1.7 ± 0.7) × 10 3 M -1 and (6.7 ± 0.2) × 10 3 M -1 for ASA and SA, respectively. Both ligands quench the fluorescence emission of ethidium bromide (Et)-ctDNA complex. The Scatchard plots indicate the non-displacement based quenching (non-intercalative binding). The circular dichroism (CD) spectra of ASA- or SA-ctDsNA complexes show the minor distortion of ctDNA structure, with no characteristic peaks for intercalation of ligands. Tm of ctDNA is decreased up to 3 °C upon ASA binding. The CD results also indicate more distortions on oligo(dG·dC) 15 structure due to the binding of both ASA and SA in comparison with oligo(dA·dT) 15. All data indicate the more affinity for SA binding with DNA minor groove in comparison with ASA which has more hydrophobic character.

Light-emitting self-assembled peptide nucleic acids exhibit both stacking interactions and Watson-Crick base pairing.

PubMed

Berger, Or; Adler-Abramovich, Lihi; Levy-Sakin, Michal; Grunwald, Assaf; Liebes-Peer, Yael; Bachar, Mor; Buzhansky, Ludmila; Mossou, Estelle; Forsyth, V Trevor; Schwartz, Tal; Ebenstein, Yuval; Frolow, Felix; Shimon, Linda J W; Patolsky, Fernando; Gazit, Ehud

2015-04-01

The two main branches of bionanotechnology involve the self-assembly of either peptides or DNA. Peptide scaffolds offer chemical versatility, architectural flexibility and structural complexity, but they lack the precise base pairing and molecular recognition available with nucleic acid assemblies. Here, inspired by the ability of aromatic dipeptides to form ordered nanostructures with unique physical properties, we explore the assembly of peptide nucleic acids (PNAs), which are short DNA mimics that have an amide backbone. All 16 combinations of the very short di-PNA building blocks were synthesized and assayed for their ability to self-associate. Only three guanine-containing di-PNAs-CG, GC and GG-could form ordered assemblies, as observed by electron microscopy, and these di-PNAs efficiently assembled into discrete architectures within a few minutes. The X-ray crystal structure of the GC di-PNA showed the occurrence of both stacking interactions and Watson-Crick base pairing. The assemblies were also found to exhibit optical properties including voltage-dependent electroluminescence and wide-range excitation-dependent fluorescence in the visible region.

Interfacial engineering of pyridinium gemini surfactants for the generation of synthetic transfection systems.

PubMed

Sharma, Vishnu D; Aifuwa, Eronmwon O; Heiney, Paul A; Ilies, Marc A

2013-09-01

Pyridinium gemini surfactants possess a soft charge, a high charge/mass ratio and a high molecular flexibility - all key parameters that recommend their use in synthetic gene delivery systems with in vitro and in vivo efficiency. In present study we generated a DNA delivery system through interfacial engineering of pyridinium gemini surfactants at the level of linker, hydrophobic chains and counterions. The self-assembling of the pyridinium amphiphiles and the physicochemical properties of the resultant supra-molecular assemblies were studied in bulk and in solution through a combination of techniques that included DSC, X-ray diffraction, polarized microscopy, CMC, dynamic light scattering and zeta potential measurements. We assessed the impact of different structural elements and formulation parameters of these pyridinium amphiphiles on their DNA compaction properties, transfection efficiency, cytotoxicity, in a complete structure-activity relationship study. This interfacial engineering process generated transfection systems with reduced cytotoxicity and high transfection efficiency in media containing elevated levels of serum that mimic the in vivo conditions. Copyright © 2013 Elsevier Ltd. All rights reserved.

Structural Basis for Intrinsic Thermosensing by the Master Virulence Regulator RovA of Yersinia*

PubMed Central

Quade, Nick; Mendonca, Chriselle; Herbst, Katharina; Heroven, Ann Kathrin; Ritter, Christiane; Heinz, Dirk W.; Dersch, Petra

2012-01-01

Pathogens often rely on thermosensing to adjust virulence gene expression. In yersiniae, important virulence-associated traits are under the control of the master regulator RovA, which uses a built-in thermosensor to control its activity. Thermal upshifts encountered upon host entry induce conformational changes in the RovA dimer that attenuate DNA binding and render the protein more susceptible to proteolysis. Here, we report the crystal structure of RovA in the free and DNA-bound forms and provide evidence that thermo-induced loss of RovA activity is promoted mainly by a thermosensing loop in the dimerization domain and residues in the adjacent C-terminal helix. These determinants allow partial unfolding of the regulator upon an upshift to 37 °C. This structural distortion is transmitted to the flexible DNA-binding domain of RovA. RovA contacts mainly the DNA backbone in a low-affinity binding mode, which allows the immediate release of RovA from its operator sites. We also show that SlyA, a close homolog of RovA from Salmonella with a very similar structure, is not a thermosensor and remains active and stable at 37 °C. Strikingly, changes in only three amino acids, reflecting evolutionary replacements in SlyA, result in a complete loss of the thermosensing properties of RovA and prevent degradation. In conclusion, only minor alterations can transform a thermotolerant regulator into a thermosensor that allows adjustment of virulence and fitness determinants to their thermal environment. PMID:22936808

Structural Insights into the Assembly of the Adeno-associated Virus Type 2 Rep68 Protein on the Integration Site AAVS1*

PubMed Central

Musayev, Faik N.; Zarate-Perez, Francisco; Bishop, Clayton; Burgner, John W.; Escalante, Carlos R.

2015-01-01

Adeno-associated virus (AAV) is the only eukaryotic virus with the property of establishing latency by integrating site-specifically into the human genome. The integration site known as AAVS1 is located in chromosome 19 and contains multiple GCTC repeats that are recognized by the AAV non-structural Rep proteins. These proteins are multifunctional, with an N-terminal origin-binding domain (OBD) and a helicase domain joined together by a short linker. As a first step to understand the process of site-specific integration, we proceeded to characterize the recognition and assembly of Rep68 onto the AAVS1 site. We first determined the x-ray structure of AAV-2 Rep68 OBD in complex with the AAVS1 DNA site. Specificity is achieved through the interaction of a glycine-rich loop that binds the major groove and an α-helix that interacts with a downstream minor groove on the same face of the DNA. Although the structure shows a complex with three OBD molecules bound to the AAVS1 site, we show by using analytical centrifugation and electron microscopy that the full-length Rep68 forms a heptameric complex. Moreover, we determined that a minimum of two direct repeats is required to form a stable complex and to melt DNA. Finally, we show that although the individual domains bind DNA poorly, complex assembly requires oligomerization and cooperation between its OBD, helicase, and the linker domains. PMID:26370092

High precision and high yield fabrication of dense nanoparticle arrays onto DNA origami at statistically independent binding sites

NASA Astrophysics Data System (ADS)

Takabayashi, Sadao; Klein, William P.; Onodera, Craig; Rapp, Blake; Flores-Estrada, Juan; Lindau, Elias; Snowball, Lejmarc; Sam, Joseph T.; Padilla, Jennifer E.; Lee, Jeunghoon; Knowlton, William B.; Graugnard, Elton; Yurke, Bernard; Kuang, Wan; Hughes, William L.

2014-10-01

High precision, high yield, and high density self-assembly of nanoparticles into arrays is essential for nanophotonics. Spatial deviations as small as a few nanometers can alter the properties of near-field coupled optical nanostructures. Several studies have reported assemblies of few nanoparticle structures with controlled spacing using DNA nanostructures with variable yield. Here, we report multi-tether design strategies and attachment yields for homo- and hetero-nanoparticle arrays templated by DNA origami nanotubes. Nanoparticle attachment yield via DNA hybridization is comparable with streptavidin-biotin binding. Independent of the number of binding sites, >97% site-occupation was achieved with four tethers and 99.2% site-occupation is theoretically possible with five tethers. The interparticle distance was within 2 nm of all design specifications and the nanoparticle spatial deviations decreased with interparticle spacing. Modified geometric, binomial, and trinomial distributions indicate that site-bridging, steric hindrance, and electrostatic repulsion were not dominant barriers to self-assembly and both tethers and binding sites were statistically independent at high particle densities.High precision, high yield, and high density self-assembly of nanoparticles into arrays is essential for nanophotonics. Spatial deviations as small as a few nanometers can alter the properties of near-field coupled optical nanostructures. Several studies have reported assemblies of few nanoparticle structures with controlled spacing using DNA nanostructures with variable yield. Here, we report multi-tether design strategies and attachment yields for homo- and hetero-nanoparticle arrays templated by DNA origami nanotubes. Nanoparticle attachment yield via DNA hybridization is comparable with streptavidin-biotin binding. Independent of the number of binding sites, >97% site-occupation was achieved with four tethers and 99.2% site-occupation is theoretically possible with five tethers. The interparticle distance was within 2 nm of all design specifications and the nanoparticle spatial deviations decreased with interparticle spacing. Modified geometric, binomial, and trinomial distributions indicate that site-bridging, steric hindrance, and electrostatic repulsion were not dominant barriers to self-assembly and both tethers and binding sites were statistically independent at high particle densities. Electronic supplementary information (ESI) available. See DOI: 10.1039/c4nr03069a

Intrastrand triplex DNA repeats in bacteria: a source of genomic instability

PubMed Central

Holder, Isabelle T.; Wagner, Stefanie; Xiong, Peiwen; Sinn, Malte; Frickey, Tancred; Meyer, Axel; Hartig, Jörg S.

2015-01-01

Repetitive nucleic acid sequences are often prone to form secondary structures distinct from B-DNA. Prominent examples of such structures are DNA triplexes. We observed that certain intrastrand triplex motifs are highly conserved and abundant in prokaryotic genomes. A systematic search of 5246 different prokaryotic plasmids and genomes for intrastrand triplex motifs was conducted and the results summarized in the ITxF database available online at http://bioinformatics.uni-konstanz.de/utils/ITxF/. Next we investigated biophysical and biochemical properties of a particular G/C-rich triplex motif (TM) that occurs in many copies in more than 260 bacterial genomes by CD and nuclear magnetic resonance spectroscopy as well as in vivo footprinting techniques. A characterization of putative properties and functions of these unusually frequent nucleic acid motifs demonstrated that the occurrence of the TM is associated with a high degree of genomic instability. TM-containing genomic loci are significantly more rearranged among closely related Escherichia coli strains compared to control sites. In addition, we found very high frequencies of TM motifs in certain Enterobacteria and Cyanobacteria that were previously described as genetically highly diverse. In conclusion we link intrastrand triplex motifs with the induction of genomic instability. We speculate that the observed instability might be an adaptive feature of these genomes that creates variation for natural selection to act upon. PMID:26450966

Inner-shell chemical shift of DNA/RNA bases and inheritance from their parent purine and pyrimidine.

PubMed

Wang, Feng; Zhu, Quan; Ivanova, Elena

2008-11-01

Inner-shell electronic structures, properties and ionization spectra of DNA/RNA bases are studied with respect to their parent pyrimidine and purine species. Density functional theory B3LYP/aug-cc-pVTZ has been employed to produce the geometries of the bases, whereas LB94/et-pVQZ//B3LYP/aug-cc-pVTZ is used to calculate site-related Hirshfeld charges and core (vertical) ionization energies, as well as inner-shell spectra of C1s, N1s and O1s for DNA/RNA bases and their parent pyrimidine and purine species. The site-dependent variations of properties indicate the changes and inheritance of chemical environment when pyrimidine and purine become substituted. In general, although the changes are site-dependent, they are also ring-dependent. Pyrimidine bases change less significantly with respect to their parent pyrimidine than the purine bases with respect to their parent purine. Pyrimidine bases such as uracil, thymine and cytosine inherit certain properties from their parent pyrimidine, such as the Hirshfeld charge distributions and the order of core ionization energy level etc. No particular sites in the pyrimidine derivatives are engaged with a dramatic chemical shift nor with energy crossings to other sites. For the core shell spectra, the purine bases inherit very little from their parent purine, and guanine exhibits the least similarities to the parent among all the DNA/RNA bases.

Theoretical study of the Hoogsteen-Watson-Crick junctions in DNA.

PubMed

Cubero, Elena; Luque, F Javier; Orozco, Modesto

2006-02-01

A series of d (AT)(n) oligonucleotides containing mixtures of normal B-type Watson-Crick and antiparallel Hoogsteen helices have been studied using molecular dynamics simulation techniques to analyze the structural and thermodynamic impact of the junction between Watson-Crick and antiparallel Hoogsteen structures. Analysis of molecular dynamics simulations strongly suggests that for all oligonucleotides studied the antiparallel Hoogsteen appears as a reasonable conformation, only slightly less stable than the canonical B-type Watson-Crick one. The junctions between the Watson-Crick and Hoogsteen structures introduces a priori a sharp discontinuity in the helix, because the properties of each type of conformation are very well preserved in the corresponding fragments. However, and quite counterintuitively, junctions do not largely distort the duplex in structural, dynamics or energetic terms. Our results strongly support the possibility that small fragments of antiparallel Hoogsteen duplex might be embedded into large fragments of B-type Watson-Crick helices, making possible protein-DNA interactions that are specific of the antiparallel Hoogsteen conformation.

Theoretical Study of the Hoogsteen–Watson-Crick Junctions in DNA

PubMed Central

Cubero, Elena; Luque, F. Javier; Orozco, Modesto

2006-01-01

A series of d (AT)n oligonucleotides containing mixtures of normal B-type Watson-Crick and antiparallel Hoogsteen helices have been studied using molecular dynamics simulation techniques to analyze the structural and thermodynamic impact of the junction between Watson-Crick and antiparallel Hoogsteen structures. Analysis of molecular dynamics simulations strongly suggests that for all oligonucleotides studied the antiparallel Hoogsteen appears as a reasonable conformation, only slightly less stable than the canonical B-type Watson-Crick one. The junctions between the Watson-Crick and Hoogsteen structures introduces a priori a sharp discontinuity in the helix, because the properties of each type of conformation are very well preserved in the corresponding fragments. However, and quite counterintuitively, junctions do not largely distort the duplex in structural, dynamics or energetic terms. Our results strongly support the possibility that small fragments of antiparallel Hoogsteen duplex might be embedded into large fragments of B-type Watson-Crick helices, making possible protein-DNA interactions that are specific of the antiparallel Hoogsteen conformation. PMID:16287814

Dynamic investigation of DNA bending and wrapping by type II topoisomerases

NASA Astrophysics Data System (ADS)

Shao, Qing; Finzi, Laura; Dunlap, David

2009-11-01

Type II topoisomerases catalyze DNA decatenation and unwinding which is crucial for cell division, and therefore type II topoisomerases are some of the main targets of anti-cancer drugs. A recent crystal structure shows that, during the catalytic cycle, a yeast type II topoimerase can bend a 10 base pair DNA segment by up to 150 degrees. Bacterial gyrase, another type II topoisomerase, can wrap DNA into a tight 180 degree turn. Bending a stiff polymer like DNA requires considerable energy and could represent the rate limiting step in the catalytic (topological) cycle. Using modified deoxyribonucleotides in PCR reactions, stiffer DNA fragments have been produced and used as substrates for topoisomerase II-mediated relaxation of plectonemes introduced in single molecules using magnetic tweezers. The wrapping ability of gyrase decreases for diamino-purine-substituted DNA in which every base pair has three hydrogen-bonds. The overall rate of relaxation of plectonemes by recombinant human topoisomerase II alpha also decreases. These results reveal the dynamic properties of DNA bending and wrapping by type II topisomerases and suggest that A:T base pair melting is a rate determining step for bending and wrapping.

Recognition of RNA by amide modified backbone nucleic acids: molecular dynamics simulations of DNA-RNA hybrids in aqueous solution.

PubMed

Nina, Mafalda; Fonné-Pfister, Raymonde; Beaudegnies, Renaud; Chekatt, Habiba; Jung, Pierre M J; Murphy-Kessabi, Fiona; De Mesmaeker, Alain; Wendeborn, Sebastian

2005-04-27

Thermodynamic and structural properties of a chemically modified DNA-RNA hybrid in which a phosphodiester linkage is replaced by a neutral amide-3 linkage (3'-CH(2)-CONH-5') were investigated using UV melting experiments, molecular dynamics simulations in explicit water, and continuum solvent models. van't Hoff analysis of the experimental UV melting curves suggests that the significant increase of the thermodynamic stability of a 15-mer DNA-RNA with seven alternated amide-3 modifications (+11 degrees C) is mainly due to an increased binding enthalpy. To further evaluate the origin in the observed affinities differences, the electrostatic contribution to the binding free energy was calculated by solving the Poisson-Boltzmann equation numerically. The nonelectrostatic contribution was estimated as the product of a hydrophobic surface tension coefficient and the surface area that is buried upon double strand formation. Structures were taken from 10 ns molecular dynamics simulations computed in a consistent fashion using explicit solvent, counterions, and the particle-mesh Ewald procedure. The present preliminary thermodynamic study suggests that the favorable binding free energy of the amide-3 DNA single strand to the complementary RNA is equally driven by electrostatic and nonpolar contributions to the binding compared to their natural analogues. In addition, molecular dynamics simulations in explicit water were performed on an amide-3 DNA single strand and the corresponding natural DNA. Results from the conformations cluster analysis of the simulated amide-3 DNA single strand ensembles suggest that the 25% of the population sampled within 10 ns has a pre-organized conformation where the sugar C3' endo pucker is favored at the 3'-flanking nucleotides. These structural and thermodynamic features contribute to the understanding of the observed increased affinities of the amide-3 DNA-RNA hybrids at the microscopic level.

Simulations Meet Experiment to Reveal New Insights into DNA Intrinsic Mechanics

PubMed Central

Ben Imeddourene, Akli; Elbahnsi, Ahmad; Guéroult, Marc; Oguey, Christophe; Foloppe, Nicolas; Hartmann, Brigitte

2015-01-01

The accurate prediction of the structure and dynamics of DNA remains a major challenge in computational biology due to the dearth of precise experimental information on DNA free in solution and limitations in the DNA force-fields underpinning the simulations. A new generation of force-fields has been developed to better represent the sequence-dependent B-DNA intrinsic mechanics, in particular with respect to the BI ↔ BII backbone equilibrium, which is essential to understand the B-DNA properties. Here, the performance of MD simulations with the newly updated force-fields Parmbsc0εζOLI and CHARMM36 was tested against a large ensemble of recent NMR data collected on four DNA dodecamers involved in nucleosome positioning. We find impressive progress towards a coherent, realistic representation of B-DNA in solution, despite residual shortcomings. This improved representation allows new and deeper interpretation of the experimental observables, including regarding the behavior of facing phosphate groups in complementary dinucleotides, and their modulation by the sequence. It also provides the opportunity to extensively revisit and refine the coupling between backbone states and inter base pair parameters, which emerges as a common theme across all the complementary dinucleotides. In sum, the global agreement between simulations and experiment reveals new aspects of intrinsic DNA mechanics, a key component of DNA-protein recognition. PMID:26657165

RPA homologs and ssDNA processing during meiotic recombination.

PubMed

Ribeiro, Jonathan; Abby, Emilie; Livera, Gabriel; Martini, Emmanuelle

2016-06-01

Meiotic homologous recombination is a specialized process that involves homologous chromosome pairing and strand exchange to guarantee proper chromosome segregation and genetic diversity. The formation and repair of DNA double-strand breaks (DSBs) during meiotic recombination differs from those during mitotic recombination in that the homologous chromosome rather than the sister chromatid is the preferred repair template. The processing of single-stranded DNA (ssDNA) formed on intermediate recombination structures is central to driving the specific outcomes of DSB repair during meiosis. Replication protein A (RPA) is the main ssDNA-binding protein complex involved in DNA metabolism. However, the existence of RPA orthologs in plants and the recent discovery of meiosis specific with OB domains (MEIOB), a widely conserved meiosis-specific RPA1 paralog, strongly suggest that multiple RPA complexes evolved and specialized to subdivide their roles during DNA metabolism. Here we review ssDNA formation and maturation during mitotic and meiotic recombination underlying the meiotic specific features. We describe and discuss the existence and properties of MEIOB and multiple RPA subunits in plants and highlight how they can provide meiosis-specific fates to ssDNA processing during homologous recombination. Understanding the functions of these RPA homologs and how they interact with the canonical RPA subunits is of major interest in the fields of meiosis and DNA repair.

Reconstitution of wild type viral DNA in simian cells transfected with early and late SV40 defective genomes.

PubMed

O'Neill, F J; Gao, Y; Xu, X

1993-11-01

The DNAs of polyomaviruses ordinarily exist as a single circular molecule of approximately 5000 base pairs. Variants of SV40, BKV and JCV have been described which contain two complementing defective DNA molecules. These defectives, which form a bipartite genome structure, contain either the viral early region or the late region. The defectives have the unique property of being able to tolerate variable sized reiterations of regulatory and terminus region sequences, and portions of the coding region. They can also exchange coding region sequences with other polyomaviruses. It has been suggested that the bipartite genome structure might be a stage in the evolution of polyomaviruses which can uniquely sustain genome and sequence diversity. However, it is not known if the regulatory and terminus region sequences are highly mutable. Also, it is not known if the bipartite genome structure is reversible and what the conditions might be which would favor restoration of the monomolecular genome structure. We addressed the first question by sequencing the reiterated regulatory and terminus regions of E- and L-SV40 DNAs. This revealed a large number of mutations in the regulatory regions of the defective genomes, including deletions, insertions, rearrangements and base substitutions. We also detected insertions and base substitutions in the T-antigen gene. We addressed the second question by introducing into permissive simian cells, E- and L-SV40 genomes which had been engineered to contain only a single regulatory region. Analysis of viral DNA from transfected cells demonstrated recombined genomes containing a wild type monomolecular DNA structure. However, the complete defectives, containing reiterated regulatory regions, could often compete away the wild type genomes. The recombinant monomolecular genomes were isolated, cloned and found to be infectious. All of the DNA alterations identified in one of the regulatory regions of E-SV40 DNA were present in the recombinant monomolecular genomes. These and other findings indicate that the bipartite genome state can sustain many mutations which wtSV40 cannot directly sustain. However, the mutations can later be introduced into the wild type genomes when the E- and L-SV40 DNAs recombine to generate a new monomolecular genome structure.

Detection of alpha particles using DNA/Al Schottky junctions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Al-Ta'ii, Hassan Maktuff Jaber, E-mail: hassankirkukly@gmail.com, E-mail: vengadeshp@um.edu.my; Department of Physics, Faculty of Science, University of Al-Muthana, Al-Muthana 66001; Periasamy, Vengadesh, E-mail: hassankirkukly@gmail.com, E-mail: vengadeshp@um.edu.my

2015-09-21

Deoxyribonucleic acid or DNA can be utilized in an organic-metallic rectifying structure to detect radiation, especially alpha particles. This has become much more important in recent years due to crucial environmental detection needs in both peace and war. In this work, we fabricated an aluminum (Al)/DNA/Al structure and generated current–voltage characteristics upon exposure to alpha radiation. Two models were utilized to investigate these current profiles; the standard conventional thermionic emission model and Cheung and Cheung's method. Using these models, the barrier height, Richardson constant, ideality factor and series resistance of the metal-DNA-metal structure were analyzed in real time. The barriermore » height, Φ value calculated using the conventional method for non-radiated structure was 0.7149 eV, increasing to 0.7367 eV after 4 min of radiation. Barrier height values were observed to increase after 20, 30 and 40 min of radiation, except for 6, 8, and 10 min, which registered a decrease of about 0.67 eV. This was in comparison using Cheung and Cheung's method, which registered 0.6983 eV and 0.7528 eV for the non-radiated and 2 min of radiation, respectively. The barrier height values, meanwhile, were observed to decrease after 4 (0.61 eV) to 40 min (0.6945 eV). The study shows that conventional thermionic emission model could be practically utilized for estimating the diode parameters including the effect of series resistance. These changes in the electronic properties of the Al/DNA/Al junctions could therefore be utilized in the manufacture of sensitive alpha particle sensors.« less

Structure, function, and tethering of DNA-binding domains in σ 54 transcriptional activators

DOE PAGES

Vidangos, Natasha; Maris, Ann E.; Young, Anisa; ...

2013-07-02

In this paper, we compare the structure, activity, and linkage of DNA-binding domains (DBDs) from σ 54 transcriptional activators and discuss how the properties of the DBDs and the linker to the neighboring domain are affected by the overall properties and requirements of the full proteins. These transcriptional activators bind upstream of specific promoters that utilize σ 54-polymerase. Upon receiving a signal the activators assemble into hexamers, which then, through adenosine triphosphate (ATP) hydrolysis, drive a conformational change in polymerase that enables transcription initiation. We present structures of the DBDs of activators nitrogen regulatory protein C 1 (NtrC1) and Nif-likemore » homolog 2 (Nlh2) from the thermophile Aquifex aeolicus. The structures of these domains and their relationship to other parts of the activators are discussed. These structures are compared with previously determined structures of the DBDs of NtrC4, NtrC, ZraR, and factor for inversion stimulation. The N-terminal linkers that connect the DBDs to the central domains in NtrC1 and Nlh2 were studied and found to be unstructured. Additionally, a crystal structure of full-length NtrC1 was solved, but density of the DBDs was extremely weak, further indicating that the linker between ATPase and DBDs functions as a flexible tether. Flexible linking of ATPase and DBDs is likely necessary to allow assembly of the active hexameric ATPase ring. Finally, the comparison of this set of activators also shows clearly that strong dimerization of the DBD only occurs when other domains do not dimerize strongly.« less

On the path to genetic novelties: insights from programmed DNA elimination and RNA splicing.

PubMed

Catania, Francesco; Schmitz, Jürgen

2015-01-01

Understanding how genetic novelties arise is a central goal of evolutionary biology. To this end, programmed DNA elimination and RNA splicing deserve special consideration. While programmed DNA elimination reshapes genomes by eliminating chromatin during organismal development, RNA splicing rearranges genetic messages by removing intronic regions during transcription. Small RNAs help to mediate this class of sequence reorganization, which is not error-free. It is this imperfection that makes programmed DNA elimination and RNA splicing excellent candidates for generating evolutionary novelties. Leveraging a number of these two processes' mechanistic and evolutionary properties, which have been uncovered over the past years, we present recently proposed models and empirical evidence for how splicing can shape the structure of protein-coding genes in eukaryotes. We also chronicle a number of intriguing similarities between the processes of programmed DNA elimination and RNA splicing, and highlight the role that the variation in the population-genetic environment may play in shaping their target sequences. © 2015 Wiley Periodicals, Inc.

Membrane Destruction and DNA Binding of Staphylococcus aureus Cells Induced by Carvacrol and Its Combined Effect with a Pulsed Electric Field.

PubMed

Wang, Lang-Hong; Wang, Man-Sheng; Zeng, Xin-An; Zhang, Zhi-Hong; Gong, De-Ming; Huang, Yan-Bo

2016-08-17

Carvacrol (5-isopropyl-2-methylphenol, CAR) is an antibacterial ingredient that occurs naturally in the leaves of the plant Origanum vulgare. The antimicrobial mechanism of CAR against Staphylococcus aureus ATCC 43300 was investigated in the study. Analysis of the membrane fatty acids by gas chromatography-mass spectrometry (GC-MS) showed that exposure to CAR at low concentrations induced a marked increase in the level of unbranched fatty acids (from 34.90 ± 1.77% to 62.37 ± 4.26%). Moreover, CAR at higher levels severely damaged the integrity and morphologies of the S. aureus cell membrane. The DNA-binding properties of CAR were also investigated using fluorescence, circular dichroism, molecular modeling, and atomic-force microscopy. The results showed that CAR bound to DNA via the minor-groove mode, mildly perturbed the DNA secondary structure, and induced DNA molecules to be aggregated. Furthermore, a combination of CAR with a pulsed-electric field was found to exhibit strong synergistic effects on S. aureus.

An effective model of DNA like helicoidal structure: with length fluctuation nonlinearity

NASA Astrophysics Data System (ADS)

Tseytlin, Y. M.

2011-03-01

One of the natural helicoidal nanostructure, which thermomechanical features are studied carefully with the help of different mechanical models, is a DNA cell / molecule. Our study proves that the experimentally determined nonlinear fluctuations of the molecular length of DNA can be better understood by modeling the molecule as a helicoidal pretwisted nanostrip sensor with nonlinear function. The calculations presented here are in good agreement with the experimental data within 10%. Other used by many researchers mechanical models such as an elastic rod, wormlike chain (WLC), accordion bellows, or an elastic core wrapped with rigid wires do not show the possible variance nonlinearity of thermomechanical DNA molecular length fluctuations. We have found that the nonlinear variance of the length fluctuations is an intrinsic property of the micro-nano-sensors with helicoidal shape. This model allows us to estimate the persistence length and twist-stretch coupling of a DNA molecule as well. It also shows the molecule's overwinding possibility at initial stretching with correct numerical representation.

Structural insights into the functions of the FANCM-FAAP24 complex in DNA repair

PubMed Central

Yang, Hui; Zhang, Tianlong; Tao, Ye; Wang, Fang; Tong, Liang; Ding, Jianping

2013-01-01

Fanconi anemia (FA) is a genetically heterogeneous disorder associated with deficiencies in the FA complementation group network. FA complementation group M (FANCM) and FA-associated protein 24 kDa (FAAP24) form a stable complex to anchor the FA core complex to chromatin in repairing DNA interstrand crosslinks. Here, we report the first crystal structure of the C-terminal segment of FANCM in complex with FAAP24. The C-terminal segment of FANCM and FAAP24 both consist of a nuclease domain at the N-terminus and a tandem helix-hairpin-helix (HhH)2 domain at the C-terminus. The FANCM-FAAP24 complex exhibits a similar architecture as that of ApXPF. However, the variations of several key residues and the electrostatic property at the active-site region render a catalytically inactive nuclease domain of FANCM, accounting for the lack of nuclease activity. We also show that the first HhH motif of FAAP24 is a potential binding site for DNA, which plays a critical role in targeting FANCM-FAAP24 to chromatin. These results reveal the mechanistic insights into the functions of FANCM-FAAP24 in DNA repair. PMID:24003026

Stabilising the Herpes Simplex Virus capsid by DNA packaging

NASA Astrophysics Data System (ADS)

Wuite, Gijs; Radtke, Kerstin; Sodeik, Beate; Roos, Wouter

2009-03-01

Three different types of Herpes Simplex Virus type 1 (HSV-1) nuclear capsids can be distinguished, A, B and C capsids. These capsids types are, respectively, empty, contain scaffold proteins, or hold DNA. We investigate the physical properties of these three capsids by combining biochemical and nanoindentation techniques. Atomic Force Microscopy (AFM) experiments show that A and C capsids are mechanically indistinguishable whereas B capsids already break at much lower forces. By extracting the pentamers with 2.0 M GuHCl or 6.0 M Urea we demonstrate an increased flexibility of all three capsid types. Remarkably, the breaking force of the B capsids without pentamers does not change, while the modified A and C capsids show a large drop in their breaking force to approximately the value of the B capsids. This result indicates that upon DNA packaging a structural change at or near the pentamers occurs which mechanically reinforces the capsids structure. The reported binding of proteins UL17/UL25 to the pentamers of the A and C capsids seems the most likely candidate for such capsids strengthening. Finally, the data supports the view that initiation of DNA packaging triggers the maturation of HSV-1 capsids.

Wavelet analysis of frequency chaos game signal: a time-frequency signature of the C. elegans DNA.

PubMed

Messaoudi, Imen; Oueslati, Afef Elloumi; Lachiri, Zied

2014-12-01

Challenging tasks are encountered in the field of bioinformatics. The choice of the genomic sequence's mapping technique is one the most fastidious tasks. It shows that a judicious choice would serve in examining periodic patterns distribution that concord with the underlying structure of genomes. Despite that, searching for a coding technique that can highlight all the information contained in the DNA has not yet attracted the attention it deserves. In this paper, we propose a new mapping technique based on the chaos game theory that we call the frequency chaos game signal (FCGS). The particularity of the FCGS coding resides in exploiting the statistical properties of the genomic sequence itself. This may reflect important structural and organizational features of DNA. To prove the usefulness of the FCGS approach in the detection of different local periodic patterns, we use the wavelet analysis because it provides access to information that can be obscured by other time-frequency methods such as the Fourier analysis. Thus, we apply the continuous wavelet transform (CWT) with the complex Morlet wavelet as a mother wavelet function. Scalograms that relate to the organism Caenorhabditis elegans (C. elegans) exhibit a multitude of periodic organization of specific DNA sequences.

Photo-degradation of CT-DNA with a series of carbothioamide ruthenium (II) complexes - Synthesis and structural analysis

NASA Astrophysics Data System (ADS)

Muthuraj, V.; Umadevi, M.

2018-04-01

The present research article is related with the method of preparation, structure and spectroscopic properties of a series of carbothioamide ruthenium (II) complexes with N and S donor ligands namely, 2-((6-chloro-4-oxo-4H-chromen-3-yl)methylene) hydrazine carbothioamide (ClChrTs)/2-((6-methoxy-4-oxo-4H-chromen-3-yl)methylene)hydrazine carbothioamide (MeOChrTS). The synthesized complexes were characterized by several techniques using analytical methods as well as by spectral techniques such as FT-IR, 1HNMR, 13CNMR, ESI mass and thermogravimetry/differential thermal analysis (TG-DTA). The IR spectra shows that the ligand acts as a neutral bidentate with N and S donor atoms. The biological activity of the prepared compounds and metal complexes were tested against cell line of calf-thymus DNA via an intercalation mechanism (MCF-7). In addition, the interaction of Ru(II) complexes and its free ligands with CT-DNA were also investigated by titration with UV-Vis spectra, fluorescence spectra, and Circular dichroism studies. Results suggest that both of the two Ru(II) complexes can bind with calf-thymus DNA via an intercalation mechanism.

The Coding of Biological Information: From Nucleotide Sequence to Protein Recognition

NASA Astrophysics Data System (ADS)

Štambuk, Nikola

The paper reviews the classic results of Swanson, Dayhoff, Grantham, Blalock and Root-Bernstein, which link genetic code nucleotide patterns to the protein structure, evolution and molecular recognition. Symbolic representation of the binary addresses defining particular nucleotide and amino acid properties is discussed, with consideration of: structure and metric of the code, direct correspondence between amino acid and nucleotide information, and molecular recognition of the interacting protein motifs coded by the complementary DNA and RNA strands.

Equilibrium properties of DNA and other semiflexible polymers confined in nanochannels

NASA Astrophysics Data System (ADS)

Muralidhar, Abhiram

Recent developments in next-generation sequencing (NGS) techniques have opened the door for low-cost, high-throughput sequencing of genomes. However, these developments have also exposed the inability of NGS to track large scale genomic information, which are extremely important to understand the relationship between genotype and phenotype. Genome mapping offers a reliable way to obtain information about large-scale structural variations in a given genome. A promising variant of genome mapping involves confining single DNA molecules in nanochannels whose cross-sectional dimensions are approximately 50 nm. Despite the development and commercialization of nanochannel-based genome mapping technology, the polymer physics of DNA in confinement is only beginning to be understood. Apart from its biological relevance, DNA is also used as a model polymer in experiments by polymer physicists. Indeed, the seminal experiments by Reisner et al. (2005) of DNA confined in nanochannels of different widths revealed discrepancies with the classical theories of Odijk and de Gennes for polymer confinement. Picking up from the conclusions of the dissertation of Tree (2014), this dissertation addresses a number of key outstanding problems in the area of nanoconfined DNA. Adopting a Monte Carlo chain growth technique known as the pruned-enriched Rosenbluth method, we examine the equilibrium and near-equilibrium properties of DNA and other semiflexible polymers in nanochannel confinement. We begin by analyzing the dependence of molecular weight on various thermodynamic properties of confined semiflexible polymers. This allows us to point out the finite size effects that can occur when using low molecular weight DNA in experiments. We then analyze the statistics of backfolding and hairpin formation in the context of existing theories and discuss how our results can be used to engineer better conditions for genome mapping. Finally, we elucidate the diffusion behavior of confined semiflexible polymers by comparing and contrasting our results for asymptotically long chains with other similar studies in the literature. We expect our findings to be not only beneficial to the design of better genome mapping devices, but also to the fundamental understanding of semiflexible polymers in confinement.

Antigen discovery and delivery of subunit vaccines by nonliving bacterial ghost vectors.

PubMed

Walcher, Petra; Mayr, Ulrike B; Azimpour-Tabrizi, Chakameh; Eko, Francis O; Jechlinger, Wolfgang; Mayrhofer, Peter; Alefantis, Tim; Mujer, Cesar V; DelVecchio, Vito G; Lubitz, Werner

2004-12-01

The bacterial ghost (BG) platform system is a novel vaccine delivery system endowed with intrinsic adjuvant properties. BGs are nonliving Gram-negative bacterial cell envelopes which are devoid of their cytoplasmic contents, yet maintain their cellular morphology and antigenic structures, including bioadhesive properties. The main advantages of BGs as carriers of subunit vaccines include their ability to stimulate a high immune response and to target the carrier itself to primary antigen-presenting cells. The intrinsic adjuvant properties of BGs enhance the immune response to target antigens, including T-cell activation and mucosal immunity. Since native and foreign antigens can be carried in the envelope complex of BGs, combination vaccines with multiple antigens of diverse origin can be presented to the immune system simultaneously. Beside the capacity of BGs to function as carriers of protein antigens, they also have a high loading capacity for DNA. Thus, loading BGs with recombinant DNA takes advantage of the excellent bioavailability for DNA-based vaccines and the high expression rates of the DNA-encoded antigens in target cell types such as macrophages and dendritic cells. There are many spaces within BGs including the inner and outer membranes, the periplasmic space and the internal lumen which can carry antigens, DNA or mediators of the immune response. All can be used for subunit antigen to design new vaccine candidates with particle presentation technology. In addition, the fact that BGs can also carry piggyback large-size foreign antigen particles, increases the technologic usefulness of BGs as combination vaccines against viral and bacterial pathogens. Furthermore, the BG antigen carriers can be stored as freeze-dried preparations at room temperature for extended periods without loss of efficacy. The potency, safety and relatively low production cost of BGs offer a significant technical advantage over currently utilized vaccine technologies.

An overview on genome organization of marine organisms.

PubMed

Costantini, Maria

2015-12-01

In this review we will concentrate on some general genome features of marine organisms and their evolution, ranging from vertebrate to invertebrates until unicellular organisms. Before genome sequencing, the ultracentrifugation in CsCl led to high resolution of mammalian DNA (without seeing at the sequence). The analytical profile of human DNA showed that the vertebrate genome is a mosaic of isochores, typically megabase-size DNA segments that belong in a small number of families characterized by different GC levels. The recent availability of a number of fully sequenced genomes allowed mapping very precisely the isochores, based on DNA sequences. Since isochores are tightly linked to biological properties such as gene density, replication timing and recombination, the new level of detail provided by the isochore map helped the understanding of genome structure, function and evolution. This led the current level of knowledge and to further insights. Copyright © 2015. Published by Elsevier B.V.

Electronic properties of long DNA nanowires in dry and wet conditions

NASA Astrophysics Data System (ADS)

Mousavi, Hamze; Khodadadi, Jabbar; Grabowski, Marek

2015-11-01

The electronic behavior of the long disordered DNA nanowires in both dry and wet conditions is investigated through the band structure and density of states of a tight-binding Hamiltonian model for π-electrons of the backbone, using Green's functions approach. For a chosen set of parameters in the dry case, semiconducting behavior is reproduced. It is also shown that for sufficiently long strands, the order of the base pairs has no noticeable effect on the energy band-gap. Moreover, this semiconducting duplex shows metallic tendencies when interacting with the environment of polar molecules.

Diversity of small, single-stranded DNA viruses of invertebrates and their chaotic evolutionary past.

PubMed

Tijssen, Peter; Pénzes, Judit J; Yu, Qian; Pham, Hanh T; Bergoin, Max

2016-10-01

A wide spectrum of invertebrates is susceptible to various single-stranded DNA viruses. Their relative simplicity of replication and dependence on actively dividing cells makes them highly pathogenic for many invertebrates (Hexapoda, Decapoda, etc.). We present their taxonomical classification and describe the evolutionary relationships between various groups of invertebrate-infecting viruses, their high degree of recombination, and their relationship to viruses infecting mammals or other vertebrates. They share characteristics of the viruses within the various families, including structure of the virus particle, genome properties, and gene expression strategy. Copyright © 2016. Published by Elsevier Inc.

THE OPEP COARSE-GRAINED PROTEIN MODEL: FROM SINGLE MOLECULES, AMYLOID FORMATION, ROLE OF MACROMOLECULAR CROWDING AND HYDRODYNAMICS TO RNA/DNA COMPLEXES

PubMed Central

Sterpone, Fabio; Melchionna, Simone; Tuffery, Pierre; Pasquali, Samuela; Mousseau, Normand; Cragnolini, Tristan; Chebaro, Yassmine; Saint-Pierre, Jean-Francois; Kalimeri, Maria; Barducci, Alessandro; Laurin, Yohan; Tek, Alex; Baaden, Marc; Nguyen, Phuong Hoang; Derreumaux, Philippe

2015-01-01

The OPEP coarse-grained protein model has been applied to a wide range of applications since its first release 15 years ago. The model, which combines energetic and structural accuracy and chemical specificity, allows studying single protein properties, DNA/RNA complexes, amyloid fibril formation and protein suspensions in a crowded environment. Here we first review the current state of the model and the most exciting applications using advanced conformational sampling methods. We then present the current limitations and a perspective on the on-going developments. PMID:24759934

Identification and Characterization of the UL37 Protein of Herpes Simplex Virus Type 1 and Demonstration that it Interacts with ICP8, the Major DNA Binding Protein of Herpes Simplex Virus

DTIC Science & Technology

1992-10-20

Identification of ORFs HSV DNA binding proteins • 1 3 3 5 7 7 11 17 18 22 reps and its role in HSV replication 23 Biochemical properties . . 23...Figure 1 . 2. 3 • 4. 5. 6. 7. 8. Structural model of the herpesvirus virion Schematic diagram of HSV pathogenesis . Diagram of the main...vaccinia virus- 13. Autoradiogram of an immunoblot of HSV - 1 -infected cell proteins harvested at various times postinfec- 85 tioD probed with anti-UL42

Molecular dynamics of DNA quadruplex molecules containing inosine, 6-thioguanine and 6-thiopurine.

PubMed Central

Stefl, R; Spacková, N; Berger, I; Koca, J; Sponer, J

2001-01-01

The ability of the four-stranded guanine (G)-DNA motif to incorporate nonstandard guanine analogue bases 6-oxopurine (inosine, I), 6-thioguanine (tG), and 6-thiopurine (tI) has been investigated using large-scale molecular dynamics simulations. The simulations suggest that a G-DNA stem can incorporate inosines without any marked effect on its structure and dynamics. The all-inosine quadruplex stem d(IIII)(4) shows identical dynamical properties as d(GGGG)(4) on the nanosecond time scale, with both molecular assemblies being stabilized by monovalent cations residing in the channel of the stem. However, simulations carried out in the absence of these cations show dramatic differences in the behavior of d(GGGG)(4) and d(IIII)(4). Whereas vacant d(GGGG)(4) shows large fluctuations but does not disintegrate, vacant d(IIII)(4) is completely disrupted within the first nanosecond. This is a consequence of the lack of the H-bonds involving the N2 amino group that is not present in inosine. This indicates that formation of the inosine quadruplex could involve entirely different intermediate structures than formation of the guanosine quadruplex, and early association of cations in this process appears to be inevitable. In the simulations, the incorporation of 6-thioguanine and 6-thiopurine sharply destabilizes four-stranded G-DNA structures, in close agreement with experimental data. The main reason is the size of the thiogroup leading to considerable steric conflicts and expelling the cations out of the channel of the quadruplex stem. The G-DNA stem can accommodate a single thioguanine base with minor perturbations. Incorporation of a thioguanine quartet layer is associated with a large destabilization of the G-DNA stem whereas the all-thioguanine quadruplex immediately collapses. PMID:11159416

Unique active site promotes error-free replication opposite an 8-oxo-guanine lesion by human DNA polymerase iota

PubMed Central

Kirouac, Kevin N.; Ling, Hong

2011-01-01

The 8-oxo-guanine (8-oxo-G) lesion is the most abundant and mutagenic oxidative DNA damage existing in the genome. Due to its dual coding nature, 8-oxo-G causes most DNA polymerases to misincorporate adenine. Human Y-family DNA polymerase iota (polι) preferentially incorporates the correct cytosine nucleotide opposite 8-oxo-G. This unique specificity may contribute to polι’s biological role in cellular protection against oxidative stress. However, the structural basis of this preferential cytosine incorporation is currently unknown. Here we present four crystal structures of polι in complex with DNA containing an 8-oxo-G lesion, paired with correct dCTP or incorrect dATP, dGTP, and dTTP nucleotides. An exceptionally narrow polι active site restricts the purine bases in a syn conformation, which prevents the dual coding properties of 8-oxo-G by inhibiting syn/anti conformational equilibrium. More importantly, the 8-oxo-G base in a syn conformation is not mutagenic in polι because its Hoogsteen edge does not form a stable base pair with dATP in the narrow active site. Instead, the syn 8-oxo-G template base forms the most stable replicating base pair with correct dCTP due to its small pyrimidine base size and enhanced hydrogen bonding with the Hoogsteen edge of 8-oxo-G. In combination with site directed mutagenesis, we show that Gln59 in the finger domain specifically interacts with the additional O8 atom of the lesion base, which influences nucleotide selection, enzymatic efficiency, and replication stalling at the lesion site. Our work provides the structural mechanism of high-fidelity 8-oxo-G replication by a human DNA polymerase. PMID:21300901

Recent development of nano-materials used in DNA biosensors.

PubMed

Xu, Kai; Huang, Junran; Ye, Zunzhong; Ying, Yibin; Li, Yanbin

2009-01-01

As knowledge of the structure and function of nucleic acid molecules has increased, sequence-specific DNA detection has gained increased importance. DNA biosensors based on nucleic acid hybridization have been actively developed because of their specificity, speed, portability, and low cost. Recently, there has been considerable interest in using nano-materials for DNA biosensors. Because of their high surface-to-volume ratios and excellent biological compatibilities, nano-materials could be used to increase the amount of DNA immobilization; moreover, DNA bound to nano-materials can maintain its biological activity. Alternatively, signal amplification by labeling a targeted analyte with nano-materials has also been reported for DNA biosensors in many papers. This review summarizes the applications of various nano-materials for DNA biosensors during past five years. We found that nano-materials of small sizes were advantageous as substrates for DNA attachment or as labels for signal amplification; and use of two or more types of nano-materials in the biosensors could improve their overall quality and to overcome the deficiencies of the individual nano-components. Most current DNA biosensors require the use of polymerase chain reaction (PCR) in their protocols. However, further development of nano-materials with smaller size and/or with improved biological and chemical properties would substantially enhance the accuracy, selectivity and sensitivity of DNA biosensors. Thus, DNA biosensors without PCR amplification may become a reality in the foreseeable future.

Recent Development of Nano-Materials Used in DNA Biosensors

PubMed Central

Xu, Kai; Huang, Junran; Ye, Zunzhong; Ying, Yibin; Li, Yanbin

2009-01-01

As knowledge of the structure and function of nucleic acid molecules has increased, sequence-specific DNA detection has gained increased importance. DNA biosensors based on nucleic acid hybridization have been actively developed because of their specificity, speed, portability, and low cost. Recently, there has been considerable interest in using nano-materials for DNA biosensors. Because of their high surface-to-volume ratios and excellent biological compatibilities, nano-materials could be used to increase the amount of DNA immobilization; moreover, DNA bound to nano-materials can maintain its biological activity. Alternatively, signal amplification by labeling a targeted analyte with nano-materials has also been reported for DNA biosensors in many papers. This review summarizes the applications of various nano-materials for DNA biosensors during past five years. We found that nano-materials of small sizes were advantageous as substrates for DNA attachment or as labels for signal amplification; and use of two or more types of nano-materials in the biosensors could improve their overall quality and to overcome the deficiencies of the individual nano-components. Most current DNA biosensors require the use of polymerase chain reaction (PCR) in their protocols. However, further development of nano-materials with smaller size and/or with improved biological and chemical properties would substantially enhance the accuracy, selectivity and sensitivity of DNA biosensors. Thus, DNA biosensors without PCR amplification may become a reality in the foreseeable future. PMID:22346713

Electronic structure of an anticancer drug DC81 and its interaction with DNA base pairs

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tiwari, Gargi, E-mail: gargi.tiwari@rediffmail.com; Sharma, Dipendra, E-mail: d-11sharma@rediffmail.com; Dwivedi, K. K., E-mail: dwivedikarunesh4@gmail.com

The drug, 8-Hydroxy-7-methoxy-pyrrolo-[2,1-c][1,4] benzodiazepine-5-one, commonly christened as DC81 belongs to the pyrrolo-[2,1-c][1,4]benzodiazepine (PBDs) family. It is a member of the group of naturally occurring antitumour antibiotics produced by various Streptomyces species. The antitumour activity of DC81 is attributed to its sequence specific interaction with G-C rich DNA region in particular, for Pu-G-Pu motifs. In the present paper, physico-chemical properties DC81 have been carried out using an ab-initio method, HF/6-31G(d,p) with GAMESS program. MEP, HOMO and LUMO surfaces have been scanned. Ionization potential, electron affinity, electronegativity, global hardness and softness of the drug have been calculated. Further, drug-DNA interactions have beenmore » examined using modified second order perturbation theory along with multicentred-multipole expansion technique. Results have been discussed in the light of other theoretical and experimental observations. Efforts have been made to elucidate the binding patterns and thereby biological properties of the drug.« less

SOD activity and DNA binding properties of a new symmetric porphyrin Schiff base ligand and its metal complexes.

PubMed

Çay, Sevim; Köse, Muhammet; Tümer, Ferhan; Gölcü, Ayşegül; Tümer, Mehmet

2015-12-05

4-Methoxy-2,6-bis(hydroxymethyl)phenol (1) was prepared from the reaction of 4-methoxyphenol and formaldehyde. The compound (1) was then oxidized to the 4-methoxy-2,6-diformylphenol (2) compound. Molecular structure of compound (2) was determined by X-ray diffraction method. A new symmetric porphyrin Schiff base ligand 4-methoxy-2,6-bis[5-(4-iminophenyl)-10,15,20-triphenylporphyrin]phenol (L) was prepared from the reaction of the 5-(4-aminophenyl)-10,15,20-triphenylporphyrin (TTP-NH2) and the compound (2) in the toluene solution. The metal complexes (Cu(II), Fe(III), Mn(III), Pt(II) and Zn(II)) of the ligand (L) were synthesized and characterized by the spectroscopic and analytical methods. The DNA (fish sperm FSdsDNA) binding studies of the ligand and its complexes were performed using UV-vis spectroscopy. Additionally, superoxide dismutase activities of the porphyrin Schiff base metal complexes were investigated. Additionally, electrochemical, photoluminescence and thermal properties of the compounds were investigated. Copyright © 2015 Elsevier B.V. All rights reserved.

Electronic structure of an anticancer drug DC81 and its interaction with DNA base pairs

NASA Astrophysics Data System (ADS)

Tiwari, Gargi; Sharma, Dipendra; Dwivedi, K. K.; Dwivedi, M. K.

2016-05-01

The drug, 8-Hydroxy-7-methoxy-pyrrolo-[2,1-c][1,4] benzodiazepine-5-one, commonly christened as DC81 belongs to the pyrrolo-[2,1-c][1,4]benzodiazepine (PBDs) family. It is a member of the group of naturally occurring antitumour antibiotics produced by various Streptomyces species. The antitumour activity of DC81 is attributed to its sequence specific interaction with G-C rich DNA region in particular, for Pu-G-Pu motifs. In the present paper, physico-chemical properties DC81 have been carried out using an ab-initio method, HF/6-31G(d,p) with GAMESS program. MEP, HOMO and LUMO surfaces have been scanned. Ionization potential, electron affinity, electronegativity, global hardness and softness of the drug have been calculated. Further, drug-DNA interactions have been examined using modified second order perturbation theory along with multicentred-multipole expansion technique. Results have been discussed in the light of other theoretical and experimental observations. Efforts have been made to elucidate the binding patterns and thereby biological properties of the drug.

Mitochondrial genome and epigenome: two sides of the same coin.

PubMed

D'Aquila, Patrizia; Montesanto, Alberto; Guarasci, Francesco; Passarino, Giuseppe; Bellizzi, Dina

2017-01-01

The involvement of mitochondrial content, structure and function as well as of the mitochondrial genome (mtDNA) in cell biology, by participating in the main processes occurring in the cells, has been a topic of intense interest for many years. More specifically, the progressive accumulation of variations in mtDNA of post-mitotic tissues represents a major contributing factor to both physiological and pathological phenotypes. Recently, an epigenetic overlay on mtDNA genetics is emerging, as demonstrated by the implication of the mitochondrial genome in the regulation of the intracellular epigenetic landscape being itself object of epigenetic modifications. Indeed, in vitro and population studies strongly suggest that, similarly to nuclear DNA, also mtDNA is subject to methylation and hydroxymethylation. It follows that the mitochondrial-nucleus cross talk and mitochondrial retrograde signaling in cellular properties require a concerted functional cooperation between genetic and epigenetic changes. The present paper aims to review the current advances in mitochondrial epigenetics studies and the increasing indication of mtDNA methylation status as an attractive biomarker for peculiar pathological phenotypes and environmental exposure.

Characterization and electrochemical response of DNA functionalized 2nm gold nanoparticles confined in a nanochannel array.

PubMed

Peinetti, Ana S; Ceretti, Helena; Mizrahi, Martín; González, Graciela A; Ramírez, Silvana A; Requejo, Félix G; Montserrat, Javier M; Battaglini, Fernando

2018-06-01

Polyvalent gold nanoparticle oligonucleotide conjugates are subject of intense research. Even though 2nm diameter AuNPs have been previously modified with DNA, little is known about their structure and electrochemical behavior. In this work, we examine the influence of different surface modification strategies on the interplay between the meso-organization and the molecular recognition properties of a 27-mer DNA strand. This DNA strand is functionalized with different sulfur-containing moieties and immobilized on 2nm gold nanoparticles confined on a nanoporous alumina, working the whole system as an electrode array. Surface coverages were determined by EXAFS and the performance as recognition elements for impedance-based sensors is evaluated. Our results prove that low DNA coverages on the confined nanoparticles prompt to a more sensitive response, showing the relevance in avoiding the DNA strand overcrowding. The system was able to determine a concentration as low as 100pM of the complementary strand, thus introducing the foundations for the construction of label-free genosensors at the nanometer scale. Copyright © 2018 Elsevier B.V. All rights reserved.

Mechanisms generating long range correlation in nucleotide composition of the Borrelia Burgdorferi genome

NASA Astrophysics Data System (ADS)

Mackiewicz, P.; Gierlik, A.; Kowalczuk, M.; Szczepanik, D.; Dudek, M. R.; Cebrat, S.

1999-12-01

We have analysed protein coding and intergenic sequences in the Borrelia burgdorferi (the Lyme disease bacterium) genome using different kinds of DNA walks. Genes occupying the leading strand of DNA have significantly different nucleotide composition from genes occupying the lagging strand. Nucleotide compositional bias of the two DNA strands reflects the aminoacid composition of proteins. 96% of genes coding for ribosomal proteins lie on the leading DNA strand, which suggests that the positions of these as well as other genes are non-random. In the B. burgdorferi genome, the asymmetry in intergenic DNA sequences is lower than the asymmetry in the third positions in codons. All these characters of the B. burgdorferi genome suggest that both replication-associated mutational pressure and recombination mechanisms have established the specific structure of the genome and now any recombination leading to inversion of a gene in respect to the direction of replication is forbidden. This property of the genome allows us to assume that it is in a steady state, which enables us to fix some parameters for simulations of DNA evolution.

Detection of damaged DNA bases by DNA glycosylase enzymes.

PubMed

Friedman, Joshua I; Stivers, James T

2010-06-22

A fundamental and shared process in all forms of life is the use of DNA glycosylase enzymes to excise rare damaged bases from genomic DNA. Without such enzymes, the highly ordered primary sequences of genes would rapidly deteriorate. Recent structural and biophysical studies are beginning to reveal a fascinating multistep mechanism for damaged base detection that begins with short-range sliding of the glycosylase along the DNA chain in a distinct conformation we call the search complex (SC). Sliding is frequently punctuated by the formation of a transient "interrogation" complex (IC) where the enzyme extrahelically inspects both normal and damaged bases in an exosite pocket that is distant from the active site. When normal bases are presented in the exosite, the IC rapidly collapses back to the SC, while a damaged base will efficiently partition forward into the active site to form the catalytically competent excision complex (EC). Here we review the unique problems associated with enzymatic detection of rare damaged DNA bases in the genome and emphasize how each complex must have specific dynamic properties that are tuned to optimize the rate and efficiency of damage site location.

Directed evolution of glutathione transferases towards a selective glutathione-binding site and improved oxidative stability.

PubMed

Axarli, Irine; Muleta, Abdi W; Chronopoulou, Evangelia G; Papageorgiou, Anastassios C; Labrou, Nikolaos E

2017-01-01

Glutathione transferases (GSTs) are a family of detoxification enzymes that catalyze the conjugation of glutathione (GSH) to electrophilic compounds. A library of alpha class GSTs was constructed by DNA shuffling using the DNA encoding the human glutathione transferase A1-1 (hGSTA1-1) and the rat glutathione transferase A1-1 (rGSTA1-1). Activity screening of the library allowed the selection of a chimeric enzyme variant (GSTD4) that displayed high affinity towards GSH and GSH-Sepharose affinity adsorbent, higher k cat /K m and improved thermal stability, compared to the parent enzymes. The crystal structures of the GSTD4 enzyme in free form and in complex with GSH were determined to 1.6Šand 2.3Šresolution, respectively. Analysis of the GSTD4 structure showed subtle conformational changes in the GSH-binding site and in electron-sharing network that may contribute to the increased GSH affinity. The shuffled variant GSTD4 was further optimized for improved oxidative stability employing site-saturation mutagenesis. The Cys112Ser mutation confers optimal oxidative stability and kinetic properties in the GSTD4 enzyme. DNA shuffling allowed the creation of a chimeric enzyme variant with improved properties, compared to the parent enzymes. X-ray crystallography shed light on how recombination of a specific segment from homologous GSTA1-1 together with point mutations gives rise to a new functionally competent enzyme with improved binding, catalytic properties and stability. Such an engineered GST would be useful in biotechnology as affinity tool in affinity chromatography as well as a biocatalytic matrix for the construction of biochips or enzyme biosensors. Copyright © 2016 Elsevier B.V. All rights reserved.

Internal-Modified Dithiol DNA–Directed Au Nanoassemblies: Geometrically Controlled Self–Assembly and Quantitative Surface–Enhanced Raman Scattering Properties

PubMed Central

Yan, Yuan; Shan, Hangyong; Li, Min; Chen, Shu; Liu, Jianyu; Cheng, Yanfang; Ye, Cui; Yang, Zhilin; Lai, Xuandi; Hu, Jianqiang

2015-01-01

In this work, a hierarchical DNA–directed self–assembly strategy to construct structure–controlled Au nanoassemblies (NAs) has been demonstrated by conjugating Au nanoparticles (NPs) with internal–modified dithiol single-strand DNA (ssDNA) (Au–B–A or A–B–Au–B–A). It is found that the dithiol–ssDNA–modified Au NPs and molecule quantity of thiol–modified ssDNA grafted to Au NPs play critical roles in the assembly of geometrically controlled Au NAs. Through matching Au–DNA self–assembly units, geometrical structures of the Au NAs can be tailored from one–dimensional (1D) to quasi–2D and 2D. Au–B–A conjugates readily give 1D and quasi–2D Au NAs while 2D Au NAs can be formed by A–B–Au–B–A building blocks. Surface-enhanced Raman scattering (SERS) measurements and 3D finite–difference time domain (3D-FDTD) calculation results indicate that the geometrically controllable Au NAs have regular and linearly “hot spots”–number–depended SERS properties. For a certain number of NPs, the number of “hot spots” and accordingly enhancement factor of Au NAs can be quantitatively evaluated, which open a new avenue for quantitative analysis based on SERS technique. PMID:26581251

Noncanonical substrate preference of lambda exonuclease for 5'-nonphosphate-ended dsDNA and a mismatch-induced acceleration effect on the enzymatic reaction.

PubMed

Wu, Tongbo; Yang, Yufei; Chen, Wei; Wang, Jiayu; Yang, Ziyu; Wang, Shenlin; Xiao, Xianjin; Li, Mengyuan; Zhao, Meiping

2018-04-06

Lambda exonuclease (λ exo) plays an important role in the resection of DNA ends for DNA repair. Currently, it is also a widely used enzymatic tool in genetic engineering, DNA-binding protein mapping, nanopore sequencing and biosensing. Herein, we disclose two noncanonical properties of this enzyme and suggest a previously undescribed hydrophobic interaction model between λ exo and DNA substrates. We demonstrate that the length of the free portion of the substrate strand in the dsDNA plays an essential role in the initiation of digestion reactions by λ exo. A dsDNA with a 5' non-phosphorylated, two-nucleotide-protruding end can be digested by λ exo with very high efficiency. Moreover, we show that when a conjugated structure is covalently attached to an internal base of the dsDNA, the presence of a single mismatched base pair at the 5' side of the modified base may significantly accelerate the process of digestion by λ exo. A detailed comparison study revealed additional π-π stacking interactions between the attached label and the amino acid residues of the enzyme. These new findings not only broaden our knowledge of the enzyme but will also be very useful for research on DNA repair and in vitro processing of nucleic acids.

Arginine homopeptides for plasmid DNA purification using monolithic supports.

PubMed

Cardoso, Sara; Sousa, Ângela; Queiroz, João A; Azzoni, Adriano R; Sousa, Fani

2018-06-15

Purification of plasmid DNA targeting therapeutic applications still presents many challenges, namely on supports and specific ligand development. Monolithic supports have emerged as interesting approaches for purifying pDNA due to its excellent mass transfer properties and higher binding capacity values. Moreover, arginine ligands were already described to establish specific and preferential interactions with pDNA. Additionally, some studies revealed the ability of arginine based cationic peptides to condense plasmid DNA, which increased lengthening can result in strongest interactions with higher binding capacities for chromatographic purposes of large molecules such as pDNA. In this work, arginine homopeptides were immobilized in monolithic supports and their performance was evaluated and compared with a single arginine monolithic column regarding supercoiled (sc) plasmid DNA purification. Specific interactions of arginine based peptides with several nucleic acids present in a clarified Escherichia coli lysate sample showed potential for the sc pDNA purification. Effectively, the immobilization of the arginine homopeptides became more functional compared with the single arginine amino acid, showing higher binding capacities, which was also reflected in the intensity of the interactions. The combination of structural versatilities of monoliths with the specificity of arginine peptides raised as a promising strategy for sc pDNA purification. Copyright © 2018 Elsevier B.V. All rights reserved.

The spectral properties of DNA and RNA macromolecules at low temperatures: fundamental and applied aspects

NASA Astrophysics Data System (ADS)

Yashchuk, Valeriy M.; Kudrya, Vladislav Yu

2017-03-01

This paper summarizes the results of studies of the spectral properties—optical absorption, fluorescence and phosphorescence—of DNA and RNA macromolecules and synthetic poly-, oligo- and mono-nucleotides, which have been carried out in our laboratory. The system of first excited singlet and triplet energy levels for DNA and RNA is evaluated using low-temperature (4.2 K-77 K) luminescent measurements. The traps of the singlet and triplet electronic excitations in these compounds are identified. An important self-protection mechanism against photo-damage of DNA and RNA by UV photons or penetrative radiation based on the capture of triplet electronic-energy excitations by the most photostable centers—in DNA, the complex formed by neighboring adenosine (A) and thymidine (T) links; in RNA, the adenosine links—is described. It is confirmed that despite similarities in the chemical and partly energy structures DNA is more stable than RNA. The spectral manifestation of the telomeres (the important functional system) in DNA macromolecules is examined. The results obtained on telomere fragments provide the possibility of finding the configuration peculiarities of the triplet excitations traps in DNA macromolecules. The resulting spreading length of the migrating singlet (l s) and triplet (l t) excitations for DNA and RNA macromolecules are evaluated.

Tangible Models and Haptic Representations Aid Learning of Molecular Biology Concepts

ERIC Educational Resources Information Center

Johannes, Kristen; Powers, Jacklyn; Couper, Lisa; Silberglitt, Matt; Davenport, Jodi

2016-01-01

Can novel 3D models help students develop a deeper understanding of core concepts in molecular biology? We adapted 3D molecular models, developed by scientists, for use in high school science classrooms. The models accurately represent the structural and functional properties of complex DNA and Virus molecules, and provide visual and haptic…

Possible prebiotic significance of polyamines in the condensation, protection, encapsulation, and biological properties of DNA

NASA Technical Reports Server (NTRS)

Baeza, I.; Ibanez, M.; Wong, C.; Chavez, P.; Gariglio, P.; Oro, J.

1991-01-01

Some properties of DNA condensed with spermidine have been compared with the properties of DNA condensed with Co3+(NH3)6 to determine whether condensation of DNA with these trivalent cations protects DNA against the action of DNase I and increases transcription and encapsulation of DNA into liposomes. It was shown that DNA condensed with Co3+(NH3)6 was resistant to the action of the endonuclease DNase I such as DNA condensed with spermidine was. However, DNA condensed with Co3+(NH3)6 was significantly less active in transcription with the E. coli RNA polymerase than DNA-spermidine condensed forms. In addition, it was demonstrated that both compacted forms of DNA were more efficiently encapsulated into neutral liposomes; however, negatively, charged liposomes were scarcely formed in the presence of DNA condensed with Co3+(NH3)6. These experiments and the well documented properties of polyamines increasing the resistance to radiations and hydrolysis of nucleic acids, as well as their biological activities, such as replication, transcription, and translation, together with the low concentration of Co3+ in the environment, lead us to propose spermidine as a plausible prebiotic DNA condensing agent rather than Co3+ and the basic proteins proposed by other authors. Then, we consider the possible role and relevance of the polyamine-nucleic acids complexes in the evolution of life.

Structural Insights Into DNA Repair by RNase T—An Exonuclease Processing 3′ End of Structured DNA in Repair Pathways

PubMed Central

Hsiao, Yu-Yuan; Fang, Woei-Horng; Lee, Chia-Chia; Chen, Yi-Ping; Yuan, Hanna S.

2014-01-01

DNA repair mechanisms are essential for preservation of genome integrity. However, it is not clear how DNA are selected and processed at broken ends by exonucleases during repair pathways. Here we show that the DnaQ-like exonuclease RNase T is critical for Escherichia coli resistance to various DNA-damaging agents and UV radiation. RNase T specifically trims the 3′ end of structured DNA, including bulge, bubble, and Y-structured DNA, and it can work with Endonuclease V to restore the deaminated base in an inosine-containing heteroduplex DNA. Crystal structure analyses further reveal how RNase T recognizes the bulge DNA by inserting a phenylalanine into the bulge, and as a result the 3′ end of blunt-end bulge DNA can be digested by RNase T. In contrast, the homodimeric RNase T interacts with the Y-structured DNA by a different binding mode via a single protomer so that the 3′ overhang of the Y-structured DNA can be trimmed closely to the duplex region. Our data suggest that RNase T likely processes bulge and bubble DNA in the Endonuclease V–dependent DNA repair, whereas it processes Y-structured DNA in UV-induced and various other DNA repair pathways. This study thus provides mechanistic insights for RNase T and thousands of DnaQ-like exonucleases in DNA 3′-end processing. PMID:24594808

The effects of structural variations of thiophene-containing Ru(II) complexes on the acid-base and DNA binding properties.

PubMed

Yuan, Cui-Li; Zhang, An-Guo; Zheng, Ze-Bo; Wang, Ke-Zhi

2013-03-01

A phenylthiophenyl-bearing Ru(II) complex of [Ru(bpy)₂(Hbptip)](PF₆)₂ {bpy = 2,2'-bipyridine, Hbptip = 2-(4-phenylthiophen-2-yl)-1H-imidazo[4,5-f][1,10]phenanthroline} was synthesized and characterized by elemental analysis, ¹H NMR spectroscopy, and electrospray ionization mass spectrometry. The ground- and excited-state acid-base properties of the complex were studied by UV-visible absorption and photoluminescence spectrophotometric pH titrations and the negative logarithm values of the ground-state acid ionization constants were derived to be pK(a1) = 1.31 ± 0.09 and pK(a2) = 5.71 ± 0.11 with the pK(a2) associated deprotonation/protonation process occurring over 3 pK(a) units more acidic than thiophenyl-free parent complex of [Ru(bpy)₂(Hpip)]²⁺ {Hpip = 2-phenyl-1H-imidazo[4,5-f][1,10]phenanthroline}. The calf thymus DNA-binding properties of [Ru(bpy)₂(Hbptip)]²⁺ in Tris-HCl buffer (pH 7.1 and 50 mM NaCl) were investigated by DNA viscosities and density functional theoretical calculations as well as UV-visible and emission spectroscopy techniques of UV-visible and luminescence titrations, steady-state emission quenching by [Fe(CN)₆]⁴⁻, DNA competitive binding with ethidium bromide, DNA melting experiments, and reverse salt effects. The complex was evidenced to bind to the DNA intercalatively with binding affinity being greater than those for previously reported analogs of [Ru(bpy)₂(Hip)]²⁺, [Ru(bpy)₂(Htip)]²⁺, and [Ru(bpy)₂(Haptip)]²⁺ {Hip = 1H-imidazo[4,5-f][1,10]phenanthroline, Htip = 2-thiophenimidazo[4,5-f][1,10]phenanthroline, Haptip = 2-(5-phenylthiophen-2-yl)-1H-imidazo[4,5-f][1,10]phenanthroline}.

Modeling the mechanical properties of DNA nanostructures.

PubMed

Arbona, Jean Michel; Aimé, Jean-Pierre; Elezgaray, Juan

2012-11-01

We discuss generalizations of a previously published coarse-grained description [Mergell et al., Phys. Rev. E 68, 021911 (2003)] of double stranded DNA (dsDNA). The model is defined at the base-pair level and includes the electrostatic repulsion between neighbor helices. We show that the model reproduces mechanical and elastic properties of several DNA nanostructures (DNA origamis). We also show that electrostatic interactions are necessary to reproduce atomic force microscopy measurements on planar DNA origamis.

Speeding up the self-assembly of a DNA nanodevice using a variety of polar solvents

NASA Astrophysics Data System (ADS)

Kang, Di; Duan, Ruixue; Tan, Yerpeng; Hong, Fan; Wang, Boya; Chen, Zhifei; Xu, Shaofang; Lou, Xiaoding; Wei, Wei; Yurke, Bernard; Xia, Fan

2014-11-01

The specific recognition and programmable assembly properties make DNA a potential material for nanodevices. However, the more intelligent the nanodevice is, the more complicated the structure of the nanodevice is, which limits the speed of DNA assembly. Herein, to address this problem, we investigate the performance of DNA Strand Displacement Reaction (DSDR) in a mixture of polar organic solvents and aqueous buffer and demonstrate that the organic polar solvent can speed up DNA self-assembly efficiently. Taking DSDR in 20% ethanol as an example, first we have demonstrated that the DSDR is highly accelerated in the beginning of the reaction and it can complete 60% of replacement reactions (160% enhancement compared with aqueous buffer) in the first 300 seconds. Secondly, we calculated that the ΔΔG of the DSDR in 20% ethanol (-18.2 kcal mol-1) is lower than that in pure aqueous buffer (-32.6 kcal mol-1), while the activation energy is lowered by introducing ethanol. Finally, we proved that the DSDR on the electrode surface can also be accelerated using this simple strategy. More importantly, to test the efficacy of this approach in nanodevices with a complicated and slow DNA self-assembly process, we apply this strategy in the hybridization chain reaction (HCR) and prove the acceleration is fairly obvious in 20% ethanol, which demonstrates the feasibility of the proposed strategy in DNA nanotechnology and DNA-based biosensors.The specific recognition and programmable assembly properties make DNA a potential material for nanodevices. However, the more intelligent the nanodevice is, the more complicated the structure of the nanodevice is, which limits the speed of DNA assembly. Herein, to address this problem, we investigate the performance of DNA Strand Displacement Reaction (DSDR) in a mixture of polar organic solvents and aqueous buffer and demonstrate that the organic polar solvent can speed up DNA self-assembly efficiently. Taking DSDR in 20% ethanol as an example, first we have demonstrated that the DSDR is highly accelerated in the beginning of the reaction and it can complete 60% of replacement reactions (160% enhancement compared with aqueous buffer) in the first 300 seconds. Secondly, we calculated that the ΔΔG of the DSDR in 20% ethanol (-18.2 kcal mol-1) is lower than that in pure aqueous buffer (-32.6 kcal mol-1), while the activation energy is lowered by introducing ethanol. Finally, we proved that the DSDR on the electrode surface can also be accelerated using this simple strategy. More importantly, to test the efficacy of this approach in nanodevices with a complicated and slow DNA self-assembly process, we apply this strategy in the hybridization chain reaction (HCR) and prove the acceleration is fairly obvious in 20% ethanol, which demonstrates the feasibility of the proposed strategy in DNA nanotechnology and DNA-based biosensors. Electronic supplementary information (ESI) available: Experimental procedures and analytical data are provided. See DOI: 10.1039/c4nr02257b

Detecting and identifying DNA via the THz backbone frequency using a metamaterial-based label-free biosensor

NASA Astrophysics Data System (ADS)

Mirzaei, Sahar; Green, Nicolas G.; Rotaru, Mihai; Pu, Suan Hui

2017-02-01

In genetic diagnostics, laboratory-based equipment generally uses analytical techniques requiring complicated and expensive fluorescent labelling of target DNA molecules. Intense research effort into, and commercial development of, Point-of-Care diagnostics and Personalized Healthcare are driving the development of simple, fast and cost-effective detection methods. One potential label-free DNA detection method uses Terahertz (THz) spectroscopy of the natural responses of DNA in metamaterial structures, which are engineered to have properties that are impossible to obtain in natural materials. This paper presents a study of the development of metamaterials based on asymmetric X-shaped resonator inclusions as a functional sensor for DNA. Gold X-shaped resonator structures with dimensions of 90/85 μm were demonstrated to produce trapped mode resonant frequency in the correct range for DNA detection. Realistic substrate materials in the form of 375 μm thick quartz were investigated, demonstrating that the non-transparent nature of the material resulted in the production of standing waves, affecting the system response, as well as requiring a reduction in scale of the resonator of 85%. As a result, the effect of introducing etched windows in the substrate material were investigated, demonstrating that increased window size significantly reduces the effect of the substrate on the system response. The device design showed a good selectivity when RNA samples were introduced to the model, demonstrating the potential for this design of device in the development of sensors capable of performing cheap and simple genetic analysis of DNA, giving label-free detection at high sensitivity.

Functional properties and structural requirements of the plasmid pMV158-encoded MobM relaxase domain.

PubMed

Fernández-López, Cris; Pluta, Radoslaw; Pérez-Luque, Rosa; Rodríguez-González, Lorena; Espinosa, Manuel; Coll, Miquel; Lorenzo-Díaz, Fabián; Boer, D Roeland

2013-07-01

A crucial element in the horizontal transfer of mobilizable and conjugative plasmids is the relaxase, a single-stranded endonuclease that nicks the origin of transfer (oriT) of the plasmid DNA. The relaxase of the pMV158 mobilizable plasmid is MobM (494 residues). In solution, MobM forms a dimer through its C-terminal domain, which is proposed to anchor the protein to the cell membrane and to participate in type 4 secretion system (T4SS) protein-protein interactions. In order to gain a deeper insight into the structural MobM requirements for efficient DNA catalysis, we studied two endonuclease domain variants that include the first 199 or 243 amino acid residues (MobMN199 and MobMN243, respectively). Our results confirmed that the two proteins behaved as monomers in solution. Interestingly, MobMN243 relaxed supercoiled DNA and cleaved single-stranded oligonucleotides harboring oriTpMV158, whereas MobMN199 was active only on supercoiled DNA. Protein stability studies using gel electrophoresis and mass spectrometry showed increased susceptibility to degradation at the domain boundary between the N- and C-terminal domains, suggesting that the domains change their relative orientation upon DNA binding. Overall, these results demonstrate that MobMN243 is capable of nicking the DNA substrate independently of its topology and that the amino acids 200 to 243 modulate substrate specificity but not the nicking activity per se. These findings suggest that these amino acids are involved in positioning the DNA for the nuclease reaction rather than in the nicking mechanism itself.

Influence of chitosan structure on the formation and stability of DNA-chitosan polyelectrolyte complexes.

PubMed

Strand, Sabina P; Danielsen, Signe; Christensen, Bjørn E; Vårum, Kjell M

2005-01-01

The interactions between DNA and chitosans varying in fractional content of acetylated units (FA), degree of polymerization (DP), and degree of ionization were investigated by several techniques, including an ethidium bromide (EtBr) fluorescence assay, gel retardation, atomic force microscopy, and dynamic and electrophoretic light scattering. The charge density of the chitosan and the number of charges per chain were found to be the dominating factors for the structure and stability of DNA-chitosan complexes. All high molecular weight chitosans condensed DNA into physically stable polyplexes; however, the properties of the complexes were strongly dependent on FA, and thereby the charge density of chitosan. By employing fully charged oligomers of constant charge density, it was shown that the complexation of DNA and stability of the polyplexes is governed by the number of cationic residues per chain. A minimum of 6-9 positive charges appeared necessary to provide interaction strength comparable to that of polycations. In contrast, further increase in the number of charges above 9 did not increase the apparent binding affinity as judged from the EtBr displacement assay. The chitosan oligomers exhibited a pH-dependent interaction with DNA, reflecting the number of ionized amino groups. The complexation of DNA and the stability of oligomer-based polyplexes became reduced above pH 7.4. Such pH-dependent dissociation of polyplexes around the physiological pH is highly relevant in gene delivery applications and might be one of the reasons for the high transfection activity of oligomer-based polyplexes observed.

Recognition of Local DNA Structures by p53 Protein

PubMed Central

Brázda, Václav; Coufal, Jan

2017-01-01

p53 plays critical roles in regulating cell cycle, apoptosis, senescence and metabolism and is commonly mutated in human cancer. These roles are achieved by interaction with other proteins, but particularly by interaction with DNA. As a transcription factor, p53 is well known to bind consensus target sequences in linear B-DNA. Recent findings indicate that p53 binds with higher affinity to target sequences that form cruciform DNA structure. Moreover, p53 binds very tightly to non-B DNA structures and local DNA structures are increasingly recognized to influence the activity of wild-type and mutant p53. Apart from cruciform structures, p53 binds to quadruplex DNA, triplex DNA, DNA loops, bulged DNA and hemicatenane DNA. In this review, we describe local DNA structures and summarize information about interactions of p53 with these structural DNA motifs. These recent data provide important insights into the complexity of the p53 pathway and the functional consequences of wild-type and mutant p53 activation in normal and tumor cells. PMID:28208646

Sequences in the intergenic spacer influence RNA Pol I transcription from the human rRNA promoter

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, W.M.; Sylvester, J.E.

1994-09-01

In most eucaryotic species, ribosomal genes are tandemly repeated about 100-5000 times per haploid genome. The 43 Kb human rDNA repeat consists of a 13 Kb coding region for the 18S, 5.8S, 28S ribosomal RNAs (rRNAs) and transcribed spacers separated by a 30 Kb intergenic spacer. For species such as frog, mouse and rat, sequences in the intergenic spacer other than the gene promoter have been shown to modulate transcription of the ribosomal gene. These sequences are spacer promoters, enhancers and the terminator for spacer transcription. We are addressing whether the human ribosomal gene promoter is similarly influenced. In-vitro transcriptionmore » run-off assays have revealed that the 4.5 kb region (CBE), directly upstream of the gene promoter, has cis-stimulation and trans-competition properties. This suggests that the CBE fragment contains an enhancer(s) for ribosomal gene transcription. Further experiments have shown that a fragment ({approximately}1.6 kb) within the CBE fragment also has trans-competition function. Deletion subclones of this region are being tested to delineate the exact sequences responsible for these modulating activities. Previous sequence analysis and functional studies have revealed that CBE contains regions of DNA capable of adopting alternative structures such as bent DNA, Z-DNA, and triple-stranded DNA. Whether these structures are required for modulating transcription remains to be determined as does the specific DNA-protein interaction involved.« less

DNA stabilization at the Bacillus subtilis PolX core—a binding model to coordinate polymerase, AP-endonuclease and 3′-5′ exonuclease activities

PubMed Central

Baños, Benito; Villar, Laurentino; Salas, Margarita; de Vega, Miguel

2012-01-01

Family X DNA polymerases (PolXs) are involved in DNA repair. Their binding to gapped DNAs relies on two conserved helix-hairpin-helix motifs, one located at the 8-kDa domain and the other at the fingers subdomain. Bacterial/archaeal PolXs have a specifically conserved third helix-hairpin-helix motif (GFGxK) at the fingers subdomain whose putative role in DNA binding had not been established. Here, mutagenesis at the corresponding residues of Bacillus subtilis PolX (PolXBs), Gly130, Gly132 and Lys134 produced enzymes with altered DNA binding properties affecting the three enzymatic activities of the protein: polymerization, located at the PolX core, 3′-5′ exonucleolysis and apurinic/apyrimidinic (AP)-endonucleolysis, placed at the so-called polymerase and histidinol phosphatase domain. Furthermore, we have changed Lys192 of PolXBs, a residue moderately conserved in the palm subdomain of bacterial PolXs and immediately preceding two catalytic aspartates of the polymerization reaction. The results point to a function of residue Lys192 in guaranteeing the right orientation of the DNA substrates at the polymerization and histidinol phosphatase active sites. The results presented here and the recently solved structures of other bacterial PolX ternary complexes lead us to propose a structural model to account for the appropriate coordination of the different catalytic activities of bacterial PolXs. PMID:22844091