Dual DNA binding mode of a turn-on red fluorescent probe thiazole coumarin.

Turn-on fluorescent probes show enhanced emission upon DNA binding, advocating their importance in imaging cellular DNA. We have probed the DNA binding mode of thiazole-coumarin (TC) conjugate, a recently reported hemicyanine-based turn-on red fluorescent probe, using a number of biophysical techniques and a series of short oligonucleotides. TC exhibited increased fluorescence anisotropy and decreased absorbance (~50%) at low [DNA]/[TC] ratio. Although the observed hypochromicity and the saturating value of [DNA base pair]:[TC] ratio is consistent with a previous study that suggested intercalation to be the DNA binding mode of TC, a distinctly different and previously unreported binding mode was observed at higher ratios of [DNA]:[TC]. With further addition of DNA, only oligonucleotides containing AnTn or (AT)n stretches showed further change-decreased hypochromicity, red shifted absorption peaks and concomitant fluorescence enhancement, saturating at about 1:1 [DNA]: [TC]. 1H-NMR chemical shift perturbation patterns and H1'-H6/H8 NOE cross-peaks of the 1:1 complex indicated minor groove binding by TC. ITC showed the 1:1 DNA binding event to be endothermic (ΔH° ~ 2 kcal/mol) and entropy driven (ΔS° ~ 32 cal/mol/K). Taken together, the experimental data suggest a dual DNA binding mode by TC. At low [DNA]/[TC] ratio, the dominant mode is intercalation. This switches to minor groove binding at higher [DNA]/[TC], only for sequences containing AnTn or (AT)n stretches. Turn-on fluorescence results only in the previously unreported minor groove bound state. Our results allow a better understanding of DNA-ligand interaction for the newly reported turn-on probe TC.

Molecular Self-Assembly of Cyclic Dipeptide Derivatives and Their Applications.

Cyclic dipeptides (CDPs) are heterocyclic 2,5-diketopiperazines with exceptional structural rigidity, enzymatic stability, and biological activity, exhibiting a substantial tendency to take part in intermolecular interactions. Strong intermolecular interactions driven by unique hydrogen bonding patterns render CDPs with a high propensity to undergo molecular self-assembly. In this Review, the aim is to provide a comprehensive summary of design strategies used to engineer the molecular self-assembly of CDPs into functional nano- and micro-architectures and molecular gels with potential applications in biomedical and materials engineering fields.

Nanoarchitectonics of Small Molecule and DNA for Ultrasensitive Detection of Mercury.

Reliable and ultrasensitive detection of mercury ions is of paramount importance for toxicology assessment, environmental protection, and human health. Herein, we present a novel optoelectronic approach based on nanoarchitectonics of small-molecule templated DNA system that consists of an adenine (A)-conjugated small organic semiconductor (BNA) and deoxyribo-oligothymidine (dTn). This mutually templated dynamic chiral coassembly system (BNAn-dTn) with tunable chiroptical, morphological, and electrical properties is tapped in to enable ultrasensitive and selective detection of inorganic and organometallic mercury in water. We observe a rapid transformation of the BNAn-dTn coassembly into a metallo-DNA duplex [dT-Hg-dT]n in the presence of mercury, which is utilized for a chiro-optical and conductivity-based rapid and subnanomolar sensitivity (≥0.1 nM, 0.02 ppb) to mercury ions in water (∼100 times lower than United States Environmental Protection Agency tolerance limit). This ultrasensitive detection of inorganic and organometallic mercury is driven by a novel chemical design principle that allows strong mercury thymine interaction. This study is anticipated to inspire the development of future templated DNA nanotechnology-based optoelectronic devices for the rapid and ultrasensitive detection of numerous other toxic analytes.

Imidazolyl-Naphthalenediimide-Based Threading Intercalators of DNA.

Intercalation by threading is anticipated to feature in DNA-binding molecules for developing DNA-targeted diagnostics and therapeutics. We investigated the role of an imidazolyl moiety in threading intercalators of DNA by employing a number of imidazolyl-naphthalenediimide conjugates. Threading intercalation was studied by UV spectroscopy, competitive binding fluorescent dye displacement, circular dichroism, isothermal calorimetry, and computational analysis. NIm6 was found to be a strong candidate, with good half-life, as revealed by dissociation kinetic analysis. Computational studies supported intercalation of the naphthalene core between base pairs and binding of the imidazolyl moieties in the adjacent grooves (threading mechanism) through electrostatic and hydrogen-bonding interactions. The interaction of the positively charged imidazolium moieties with the negatively charged phosphate backbone of DNA contributed to the favorable enthalpy change, as revealed by the experimental and computational data. Threading intercalation by NIm6 caused significant retardation of DNA in an electrophoretic mobility shift assay. The biological significance of potent imidazolyl naphthalenediimide conjugates was demonstrated by the inhibition of topo- isomerase I activity and cytotoxicity against HeLa cells.

Surface-Functionalized Silk Fibroin Films as a Platform To Guide Neuron-like Differentiation of Human Mesenchymal Stem Cells.

Surface interactions at the biomaterial-cellular interface determine the proliferation and differentiation of stem cells. Manipulating such interactions through the surface chemistry of scaffolds renders control over directed stem cell differentiation into the cell lineage of interest. This approach is of central importance for stem cell-based tissue engineering and regenerative therapy applications. In the present study, silk fibroin films (SFFs) decorated with integrin-binding laminin peptide motifs (YIGSR and GYIGSR) were prepared and employed for in vitro adult stem cell-based neural tissue engineering applications. Functionalization of SFFs with short peptides showcased the peptide sequence and nature of functionalization-dependent differentiation of bone marrow-derived human mesenchymal stem cells (hMSCs). Intriguingly, covalently functionalized SFFs with GYIGSR hexapeptide (CL2-SFF) supported hMSC proliferation and maintenance in an undifferentiated pluripotent state and directed the differentiation of hMSCs into neuron-like cells in the presence of a biochemical cue, on-demand. The observed morphological changes were further corroborated by the up-regulation of neuronal-specific marker gene expression (MAP2, TUBB3, NEFL), confirmed through semiquantitative reverse-transcription polymerase chain reaction (RT-PCR) analysis. The enhanced proliferation and on-demand directed differentiation of adult stem cells (hMSCs) by the use of an economically viable short recognition peptide (GYIGSR), as opposed to the integrin recognition protein laminin, establishes the potential of SFFs for neural tissue engineering and regenerative therapy applications.

Natural Tripeptide-Based Inhibitor of Multifaceted Amyloid ß Toxicity.

Accumulation of amyloid beta (Aß) peptide and its aggregates in the human brain is considered as one of the hallmarks of Alzheimer's disease (AD). The polymorphic oligomers and fully grown fibrillar aggregates of Aß exhibit different levels of neuronal toxicity. Moreover, aggregation of Aß in the presence of redox-active metal ions like Cu(2+) is responsible for the additional trait of cellular toxicity induced by the generation of reactive oxygen species (ROS). Herein, a multifunctional peptidomimetic inhibitor (P6) has been presented, based on a naturally occurring metal chelating tripeptide (GHK) and the inhibitor of Aß aggregation. It was shown by employing various biophysical studies that P6 interact with Aß and prevent the formation of toxic Aß forms like oligomeric species and fibrillar aggregates. Further, P6 successfully sequestered Cu(2+) from the Aß-Cu(2+) complex and maintained it in a redox-dormant state to prevent the generation of ROS. P6 inhibited membrane disruption by Aß oligomers and efficiently prevented DNA damage caused by the Aß-Cu(2+) complex. PC12 cells were rescued from multifaceted Aß toxicity when treated with P6, and the amount of ROS generated in cells was reduced. These attributes make P6 a potential therapeutic candidate to ameliorate the multifaceted Aß toxicity in AD.

Multi-Stimuli-Responsive Charge-Transfer Hydrogel for Room-Temperature Organic Ferroelectric Thin-Film Devices.

The possibility of designing programmable thin-film supramolecular structures with spontaneous polarization widens the utility of facile supramolecular chemistry. Although a range of low molecular mass molecular single crystals has been shown to exhibit ferroelectric polarization, demonstration of stimuli-responsive, thin-film, solution-processable supramolecular ferroelectric materials is rare. We introduce aromatic π-electron donor-acceptor molecular systems responsive to multiple stimuli that undergo supramolecular chiral mixed-stack charge-transfer (CT) coassembly through the tweezer-inclusion-sandwich process supported by hydrogen-bonding interactions. The structural synergy originating from hydrogen-bonding and chiral CT interactions resulted in the development of spontaneous unidirectional macroscopic polarization in the crystalline nanofibrous hydrogel network, under ambient conditions. Moreover, the tunability of these interactions with optical, mechanical, thermal, and electrical stimuli allowed the design of multistate thin-film memory devices. Our design strategy of the supramolecular motif is expected to help the development of new molecular engineering strategies for designing potentially useful smart multicomponent organic electronics.

A molecular beacon-based DNA switch for reversible pH sensing in vesicles and live cells.

In this Communication, a molecular beacon-based DNA switch (LMB) is developed as an efficient and reversible pH sensing probe. Remarkably, LMB exhibited reversible structural transition between the closed (molecular beacon) and open (A-motif) states very efficiently in synthetic vesicles and live cells without the need for any transfection agents.

Pigmented Silk Nanofibrous Composite for Skeletal Muscle Tissue Engineering.

Skeletal muscle tissue engineering (SMTE) employs designed biomaterial scaffolds for promoting myogenic differentiation of myoblasts to functional myotubes. Oxidative stress plays a significant role in the biocompatibility of biomaterials as well as in the fate of myoblasts during myogenesis and is also associated with pathological conditions such as myotonic dystrophy. The inherent electrical excitability of muscle cells inspired the use of electroactive scaffolds for SMTE. Conducting polymers attracted the attention of researchers for their use in muscle tissue engineering. However, poor biocompatibility, biodegradability and development of oxidative stress associated immunogenic response limits the extensive use of synthetic conducting polymers for SMTE. In order to address the limitations of synthetic polymers, intrinsically electroactive and antioxidant silk fibroin/melanin composite films and electrospun fiber mats were fabricated and evaluated as scaffolds for promoting myogenesis in vitro. Melanin incorporation modulated the thermal stability, electrical conductivity of scaffolds, fiber alignment in electrospun mats and imparted good antioxidant properties to the scaffolds. The composite electrospun scaffolds promoted myoblast assembly and differentiation into uniformly aligned high aspect ratio myotubes. The results highlight the significance of scaffold topography along with conductivity in promoting myogenesis and the potential application of silk nanofibrous composite as electoractive platform for SMTE.

A High Affinity Red Fluorescence and Colorimetric Probe for Amyloid ß Aggregates.

A major challenge in the Alzheimer's disease (AD) is its timely diagnosis. Amyloid ß (Aß) aggregates have been proposed as the most viable biomarker for the diagnosis of AD. Here, we demonstrate hemicyanine-based benzothiazole-coumarin (TC) as a potential probe for the detection of highly toxic Aß42 aggregates through switch-on, enhanced (~30 fold) red fluorescence (Emax = 654 nm) and characteristic colorimetric (light red to purple) optical outputs. Interestingly, TC exhibits selectivity towards Aß42 fibrils compared to other abnormal protein aggregates. TC probe show nanomolar binding affinity (Ka = 1.72 × 10(7) M(-1)) towards Aß42 aggregates and also displace ThT bound to Aß42 fibrils due to its high binding affinity. The Aß42 fibril-specific red-shift in the absorption spectra of TC responsible for the observed colorimetric optical output has been attributed to micro-environment change around the probe from hydrophilic-like to hydrophobic-like nature. The binding site, binding energy and changes in optical properties observed for TC upon interaction with Aß42 fibrils have been further validated by molecular docking and time dependent density functional theory studies.

Molecular Architectonics of Naphthalenediimides for Efficient Structure-Property Correlation.

We present a bioinspired design strategy to effectively tailor the assembly of naphthalenediimides (NDIs) into a wide variety of architectures by functionalizing with amino acid derivatives. This bioinspired process of custom designing and engineering molecular assemblies is termed "bioinspired architectonics". By employing minute structural mutations in the form of α-substituents of amino acids, we successfully engineered molecular assembly of NDIs into zero-dimensional (0D, spheres), one-dimensional (1D, fibers), and two-dimensional (2D, sheets) architectures. The 2D sheets of phenylalanine methylester appended NDI 1 showed remarkable bulk electron mobility of up to 1 cm(2) V(-1)s(-1). With the aid of photophysical, diffraction, and microscopy techniques we rationalize the effect of molecular structure with their ordering and electronic properties in an effort to find structure-property correlations via a bioinspired modular approach.

Stimuli-responsive colorimetric and NIR fluorescence combination probe for selective reporting of cellular hydrogen peroxide.

Hydrogen peroxide (H2O2) is a key reactive oxygen species and a messenger in cellular signal transduction apart from playing a vital role in many biological processes in living organisms. In this article, we present phenyl boronic acid-functionalized quinone-cyanine (QCy-BA) in combination with AT-rich DNA (exogenous or endogenous cellular DNA), i.e., QCy-BA⊂DNA as a stimuli-responsive NIR fluorescence probe for measuring in vitro levels of H2O2. In response to cellular H2O2 stimulus, QCy-BA converts into QCy-DT, a one-donor-two-acceptor (D2A) system that exhibits switch-on NIR fluorescence upon binding to the DNA minor groove. Fluorescence studies on the combination probe QCy-BA⊂DNA showed strong NIR fluorescence selectively in the presence of H2O2. Furthermore, glucose oxidase (GOx) assay confirmed the high efficiency of the combination probe QCy-BA⊂DNA for probing H2O2 generated in situ through GOx-mediated glucose oxidation. Quantitative analysis through fluorescence plate reader, flow cytometry and live imaging approaches showed that QCy-BA is a promising probe to detect the normal as well as elevated levels of H2O2 produced by EGF/Nox pathways and post-genotoxic stress in both primary and senescent cells. Overall, QCy-BA, in combination with exogenous or cellular DNA, is a versatile probe to quantify and image H2O2 in normal and disease-associated cells.

Emergent Behaviors in Kinetically Controlled Dynamic Self-Assembly of Synthetic Molecular Systems.

Living systems are categorically a kinetic state of matter that exhibits complex functions and emergent behaviors. By contrast, synthetic systems are relatively simple and are typically controlled by the thermodynamic parameters. To understand this inherent difference between the biological and synthetic systems, novel approaches are of vital importance. In this regard, we have designed a three-component molecular system (a triad) by conjugating an amino acid with two functional molecules (naphthalenediimide and pyrene), which facilitates kinetically controlled self-assemblies. Herein, we describe three different molecular aggregation states of triads (entitled State I, State II, and State III) and also the dynamic pathway complexities associated with their transformations from one state to another. By meticulously employing the triads of different molecular aggregation states and the stereochemical information of the amino acid, we report emergent behaviors termed "supramolecular speciation" and "supramolecular regulation". Further, we present a hitherto unknown emergent property in a self-assembled state under the majority-rules experiment, which has been termed "super-nonlinearity". This work provides novel insights into complex synthetic systems having unprecedented functions and properties. Such emergent behaviors of synthetic triads that involve an interplay among complex interactions may find relevance in the context of prebiotic chemical evolution.

Bioinspired Reductionistic Peptide Engineering for Exceptional Mechanical Properties.

A simple solution-processing and self-assembly approach that exploits the synergistic interactions between multiple hydrogen bonded networks and aromatic interactions was utilized to synthesize molecular crystals of cyclic dipeptides (CDPs), whose molecular weights (~0.2 kDa) are nearly three orders of magnitude smaller than that of natural structural proteins (50-300 kDa). Mechanical properties of these materials, measured using the nanoindentation technique, indicate that the stiffness and strength are comparable and sometimes better than those of natural fibres. The measured mechanical responses were rationalized by recourse to the crystallographic structural analysis and intermolecular interactions in the self-assembled single crystals. With this work we highlight the significance of developing small molecule based bioinspired design strategies to emulate biomechanical properties. A particular advantage of the successfully demonstrated reductionistic strategy of the present work is its amenability for realistic industrial scale manufacturing of designer biomaterials with desired mechanical properties.

Sequence-specific recognition of DNA minor groove by an NIR-fluorescence switch-on probe and its potential applications.

In molecular biology, understanding the functional and structural aspects of DNA requires sequence-specific DNA binding probes. Especially, sequence-specific fluorescence probes offer the advantage of real-time monitoring of the conformational and structural reorganization of DNA in living cells. Herein, we designed a new class of D2A (one-donor-two-acceptor) near-infrared (NIR) fluorescence switch-on probe named quinone cyanine-dithiazole ( QCY-DT: ) based on the distinctive internal charge transfer (ICT) process for minor groove recognition of AT-rich DNA. Interestingly, QCY-DT: exhibited strong NIR-fluorescence enhancement in the presence of AT-rich DNA compared to GC-rich and single-stranded DNAs. We show sequence-specific minor groove recognition of QCY-DT: for DNA containing 5'-AATT-3' sequence over other variable (A/T)4 sequences and local nucleobase variation study around the 5'-X(AATT)Y-3' recognition sequence revealed that X = A and Y = T are the most preferable nucleobases. The live cell imaging studies confirmed mammalian cell permeability, low-toxicity and selective staining capacity of nuclear DNA without requiring RNase treatment. Further, Plasmodium falciparum with an AT-rich genome showed specific uptake with a reasonably low IC50 value (<4 µM). The ease of synthesis, large Stokes shift, sequence-specific DNA minor groove recognition with switch-on NIR-fluorescence, photostability and parasite staining with low IC50 make QCY-DT: a potential and commercially viable DNA probe.

Function and toxicity of amyloid beta and recent therapeutic interventions targeting amyloid beta in Alzheimer's disease.

Amyloidogenesis has been implicated in a broad spectrum of diseases in which amyloid protein is invariably misfolded and deposited in cells and organs. Alzheimer's disease is one of the most devastating ailments among amyloidogenesis induced dementia. The amyloid beta (Aß) peptide derived from amyloid precursor protein (APP) is misfolded and deposited as plaques in the brain, which are said to be the hallmark of Alzheimer's disease. In normal brains physiological concentration of the Aß peptide has been indicated to be involved in modulating neurogenesis and synaptic plasticity. However, excess Aß production, its aggregation and deposition deleteriously affect a large number of biologically important pathways leading to neuronal cell death. Targeting Aß production, Aß aggregation or its clearance from the brain has been an active area of research for preventing or curing AD. Our Feature Article intends to detail the aggregation mechanism, the physiological role of the Aß peptide, elaborate its toxic effects, and outline the different classes of molecules designed in the last two years to inhibit amyloidogenic APP processing, Aß oligomerization or fibrillogenesis and to modulate different pathways for active clearance of Aß from the brain.

Fluorescence reporting of G-quadruplex structures and modulating their DNAzyme activity using polyethylenimine-pyrene conjugate.

Four-stranded G-quadruplex structure is one of the most important non-canonical secondary structures of DNA formed by guanine (G)-rich sequences. G-rich DNA sequences are known to occur in the human genome, especially in the telomere 3' end and in oncogene promoters such as c-MYC and c-KIT. In this context, we designed pyrene-conjugated polyethylenimine (PEI-Py) as a fluorescence reporter for the recognition and detection of G-quadruplex structures of G-rich deoxyoligonucleotides and human telomere and gene promoter sequences, under ambient conditions. PEI-Py exhibited prominent pyrene excimer emission in the presence of G-quadruplex structures of G-rich deoxyoligonucleotides and biologically relevant DNA sequences. PEI-Py further displayed the modulation of DNAzyme activity of various G-quadruplex structures in the presence of hemin and hydrogen peroxide.

Crystallographic insight-guided nanoarchitectonics and conductivity modulation of an n-type organic semiconductor through peptide conjugation.

Crystallographic insight-guided nanoarchitectonics of peptide-conjugated naphthalene diimide (NDI) is described. In a bio-inspired approach, non-proteinogenic α-amino isobutyric acid (Aib)- and alanine (Ala)-derived peptides orchestrated the 1D achiral and 2D chiral molecular ordering of NDI, respectively, which resulted in modulation of nanoscale morphology, chiroptical and conductivity properties.

Rationally designed peptidomimetic modulators of aß toxicity in Alzheimer's disease.

Alzheimer's disease is one of the devastating illnesses mankind is facing in the 21st century. The main pathogenic event in Alzheimer's disease is believed to be the aggregation of the ß-amyloid (Aß) peptides into toxic aggregates. Molecules that interfere with this process may act as therapeutic agents for the treatment of the disease. Use of recognition unit based peptidomimetics as inhibitors are a promising approach, as they exhibit greater protease stability compared to natural peptides. Here, we present peptidomimetic inhibitors of Aß aggregation designed based on the KLVFF (P1) sequence that is known to bind Aß aggregates. We improved inhibition efficiency of P1 by introducing multiple hydrogen bond donor-acceptor moieties (thymine/barbiturate) at the N-terminal (P2 and P3), and blood serum stability by modifying the backbone by incorporating sarcosine (N-methylglycine) units at alternate positions (P4 and P5). The peptidomimetics showed moderate to good activity in both inhibition and dissolution of Aß aggregates as depicted by thioflavin assay, circular dichroism (CD) measurements and microscopy (TEM). The activity of P4 and P5 were studied in a yeast cell model showing Aß toxicity. P4 and P5 could rescue yeast cells from Aß toxicity and Aß aggregates were cleared by the process of autophagy.

Double zipper helical assembly of deoxyoligonucleotides: mutual templating and chiral imprinting to form hybrid DNA ensembles.

Herein, the conventional and unconventional hydrogen bonding potential of adenine in APA for double zipper helical assembly of deoxyoligonucleotides is demonstrated under ambient conditions. The quantum mechanical calculations supported the formation of hybrid DNA ensembles.