Microstructures of Starch Granules with Different Amylose Contents and Allomorphs as Revealed by Scattering Techniques.

In this study, as-is (ca. 12% moisture by mass) and hydrated (50% water by mass) granules of waxy potato (WP), waxy wheat (WW), waxy maize, normal maize, and high-amylose maize (HAM) starches were investigated by using small-angle neutron and X-ray scattering (SANS and SAXS), wide-angle X-ray scattering, and ultra-small-angle neutron scattering. The SANS and SAXS data were fitted using the two-phase stacking model of alternating crystalline and amorphous layers. The partial crystalline lamellar structures inside the growth rings of granules were analyzed based on the inter-lamellar distances, thicknesses of the crystalline lamellae and amorphous layers, thickness polydispersities, and water content in each type of layer. Despite having a longer average chain length of amylopectin, the WP and HAM starches, which had B-type allomorph, had a shorter inter-lamellar distance than the other three starches with A-type allomorph. The WP starch had the most uniform crystalline lamellar thickness. After hydration, the amorphous layers were expanded, resulting in an increase of inter-layer distance. The low-angle intensity upturn in SANS and SAXS was attributed to scattering from interfaces/surfaces of larger structures, such as growth rings and macroscopic granule surfaces. Data analysis methods based on model fitting and 1D correlation function were compared. The study emphasized─owing to inherent packing disorder inside granules─that a comprehensive analysis of different parameters was essential in correlating the microstructures with starch properties.

Ultracompact gas sensor with metal-organic-framework-based differential fiber-optic Fabry-Perot nanocavities.

Refractive-index (RI)-based sensing is a major optical sensing modality that can be implemented in various spectral ranges. While it has been widely used for sensing of biochemical liquids, RI-based gas sensing, particularly small-molecule gases, is challenging due to the extremely small RI change induced by gas concentration variations. We propose a RI-based ultracompact fiber-optic differential gas sensor that employs metal-organic-framework (MOF)-based dual Fabry-Perot (FP) nanocavities. A MOF is used as the FP cavity material to enhance the sensitivity as well as the selectivity to particular gas molecules. The differential sensing scheme leverages the opposite change in the cavity-length-dependent reflection of the two FP cavities, which further enhances the sensitivity compared with single FP cavity based sensing. For proof-of-concept, a fiber-optic CO2 sensor with ZIF-8-based dual FP nanocavities was fabricated. The effective footprint of the sensor was as small as 157 µm2 and the sensor showed an enhanced sensitivity of 48.5 mV/CO2Vol%, a dynamic range of 0-100 CO2Vol%, and a resolution of 0.019 CO2Vol% with 1 Hz low-pass filtering. Although the current sensor was only demonstrated for CO2 sensing, the proposed sensor concept can be used for sensing of a variety of gases when different kinds of MOFs are utilized.

Crystal Structure of a Tetrameric DNA Fold-Back Quadruplex.

DNA can adopt many structures beyond the Watson-Crick duplex. However, the bounds of DNA structural diversity and how these structures might regulate biological processes is only beginning to be understood. Here, we describe the 1.05 Å resolution crystal structure of a DNA oligonucleotide that self-associates to form a non-G-quadruplex fold-back structure. Distinct from previously described fold-back quadruplexes, two-fold-back dimers interact through noncanonical and Watson-Crick interactions to form a tetrameric assembly. These interactions include a hexad base pairing arrangement from two C-G-G base triples. The assembly is dependent on divalent cations, and the interface between the dimeric units creates a cavity in which a cation resides. This structure provides new sequence and structural contexts for the formation of fold-back quadruplexes, further highlighting the potential biological importance of this type of noncanonical DNA structure. This structure may also serve as the basis for designing new types of DNA nanoarchitectures or cation sensors based on the strong divalent cation dependence.

Dynamic Sub-10-nm Nanostructured Ultrathin Films of Sugar-Polyolefin Conjugates Thermoresponsive at Physiological Temperatures.

Spin-casting of a cellobiose-atactic polypropene (CB-aPP) conjugate (1) from a 0.1% (w/w) n-butanol/hexane solution onto highly oriented pyrolytic graphite (HOPG) and carbon-coated Si(100) spontaneously produced microphase-separated sub-10-nm nanostructured ultrathin films in the form of alternating CB and aPP lamellar domains (d = 6.60 ± 0.68 nm) that are oriented perpendicular to the substrate surface. Thermal annealing at modest temperatures (e.g., 50-100 °C), and as low as the physiologically relevant temperature of 38 °C, serves to drive a structural transition that yields a parallel stacked bilayer assembly as the thermodynamically favored nanostructure. These results establish the advantage of low molecular weight, narrow polydispersity, and amorphous, low Tg, poly(α-olefinate)s (xPAOs) as a new class of hydrophobic building block for amphiphilic materials, and sugar-PAO conjugates in particular, for the development of stimuli-responsive, nanostructured materials for technological applications at physiological temperatures.

Decoding Single Molecule Time Traces with Dynamic Disorder.

Single molecule time trajectories of biomolecules provide glimpses into complex folding landscapes that are difficult to visualize using conventional ensemble measurements. Recent experiments and theoretical analyses have highlighted dynamic disorder in certain classes of biomolecules, whose dynamic pattern of conformational transitions is affected by slower transition dynamics of internal state hidden in a low dimensional projection. A systematic means to analyze such data is, however, currently not well developed. Here we report a new algorithm-Variational Bayes-double chain Markov model (VB-DCMM)-to analyze single molecule time trajectories that display dynamic disorder. The proposed analysis employing VB-DCMM allows us to detect the presence of dynamic disorder, if any, in each trajectory, identify the number of internal states, and estimate transition rates between the internal states as well as the rates of conformational transition within each internal state. Applying VB-DCMM algorithm to single molecule FRET data of H-DNA in 100 mM-Na+ solution, followed by data clustering, we show that at least 6 kinetic paths linking 4 distinct internal states are required to correctly interpret the duplex-triplex transitions of H-DNA.

End-Group-Functionalized Poly(α-olefinates) as Non-Polar Building Blocks: Self-Assembly of Sugar-Polyolefin Hybrid Conjugates.

Living coordinative chain-transfer polymerization of α-olefins, followed by chemical functionalization of a Zn(polymeryl)2 intermediate, provides entry to end-group functionalized poly(α-olefinates) (x-PAOs) that can serve as a new class of non-polar building block with tailorable occupied volumes. Application of these x-PAOs for the synthesis and self-assembly of sugar-polyolefin hybrid conjugates demonstrate the ability to manipulate the morphology of the ultra-thin film nanostructure through variation in occupied volume of the x-PAO domain.

Dinuclear bis-propagators for the stereoselective living coordinative chain transfer polymerization of propene.

Modulation of steric interactions remote from the active sites within a series of dinuclear bis-propagators derived from racemic 2-4 was used to attenuate the rate of reversible chain transfer between active transition-metal centers and excess equivalents of inactive main-group-metal alkyl species relative to chain growth propagation, as a strategy for achieving the stereoselective living coordinative chain transfer polymerization of propene to provide isotactic stereoblock polypropene. Under identical conditions, the corresponding mononuclear propagator derived from racemic 1 produced only atactic polypropene.

Dynamic anchoring of the 3'-end of the guide strand controls the target dissociation of Argonaute-guide complex.

Argonaute (Ago) is the catalytic core of small RNA-based gene regulation. Despite plenty of mechanistic studies on Ago, the dynamical aspects and the mechanistic determinants of target mRNA binding and dissociation of Ago-guide strand remain unclear. Here, by using single-molecule fluorescence resonance energy transfer (FRET) assays and Thermus thermophilus Ago (TtAgo), we reveal that the 3'-end of the guide strand dynamically anchors at and releases from the PAZ domain of Ago, and that the 3'-end anchoring of the guide strand greatly accelerates the target dissociation by destabilizing the guide-target duplex. Our results indicate that the target binding/dissociation of Ago-guide is executed through the dynamic interplays among Ago, guide, and target.

Dynamic competition of DsrA and rpoS fragments for the proximal binding site of Hfq as a means for efficient annealing.

Hfq is a key regulator involved in multiple aspects of stress tolerance and virulence of bacteria. There has been an intriguing question as to how this RNA chaperone achieves two completely opposite functions--annealing and unwinding--for different RNA substrates. To address this question, we studied the Hfq-mediated interaction of fragments of a non-coding RNA, DsrA, with its mRNA target rpoS by using single-molecule fluorescence techniques. These experiments permitted us to observe the mechanistic steps of Hfq-mediated RNA annealing/unwinding at the single-molecule level, for the first time. Our real-time observations reveal that, even if the ring-shaped Hfq displays multiple binding sites for its interaction with RNA, the regulatory RNA and the mRNA compete for the same binding site. The competition makes the RNA-Hfq interaction dynamic and, surprisingly, increases the overall annealing efficiency by properly aligning the two RNAs. We furthermore reveal that when Hfq specifically binds to only one of the two RNAs, the unwinding process dominates over the annealing process, thus shedding a new light on the substrate selectivity for annealing or unwinding. Finally, our results demonstrate for the first time that a single Hfq hexamer is sufficient to facilitate sRNA-mRNA annealing.

Autofocusing system based on optical astigmatism analysis of single-molecule images.

Single-molecule fluorescence imaging has greatly contributed to our understanding of many bio-molecular systems. While reactions occurring in the range of several minutes can be readily studied using conventional single-molecule fluorescence microscopes, data acquisition for longer time scales is hindered by the focal drift of high numerical aperture objectives, which should be corrected in real time. Here, we developed a robust autofocusing system based on optical astigmatism analysis of single-molecule images. Compared to the previously developed methods, our approach has a merit of simplicity in that neither fiducial makers nor an additional laser-detector system is required. As a demonstration, we observed B-Z transition dynamics occurring for several hours.

Melanin pigments extracted from horsehair as antibacterial agents.

Here we present the important findings related to biologically derived pigments for potential use as antibacterial agents. Melanin biopigments extracted from Equus ferus hair exhibit a homogeneous elliptical microstructure with highly ordered semicrystalline features. Spectroscopic analysis indicates that melanin contains a high degree of redox active catechol groups, which can produce reactive oxygen species. The antibacterial activity of melanins was tested by incubating Escherichia coli and Staphylococcus aureus with melanins. The results showed 100% bacterial growth inhibition within 4 h. This finding suggests that melanin pigments may serve as naturally occurring antibacterial agents with unique redox chemistry and reactive oxygen species generation capability.

Surface Induced nanofiber growth by self-assembly of a silk-elastin-like protein polymer.

Many synthetic and natural peptides are known to self-assemble to form various nanostructures. During the self-assembling process, environmental conditions such as salt concentration, pH, temperature, and surface characteristics play a critical role by influencing intermolecular interactions, and hence the process of self-assembly. Here we studied the self-assembly of a genetically engineered protein polymer composed of silk-like and elastin-like repeats on a mica surface. Silk-elastin-like protein polymers (SELPs) consist of tandem repeats of Gly-Ala-Gly-Ala-Gly-Ser from Bombyx mori (silkworm) and Gly-Val-Gly-Val-Pro from mammalian elastin. At a very low polymer concentration of 1 mug/mL, SELPs self-assembled into nanofibrous structures on a mica surface. Examination using atomic force microscopy (AFM) and dynamic light scattering techniques showed that SELPs self-assembled into nanofibers in the presence of the mica surface but not in the bulk state. Ionic strength had a significant influence on nanofiber growth, indicating the importance of electrostatic interactions between the polymer and the mica surface. At low ionic strength, the kinetics of nanofiber growth showed that the mica surface effectively removed a lag phase by providing nucleating sites, facilitating nanofiber self-assembly of SELPs. Furthermore, self-assembly on additional substrates such as silicon and a hydrophobic pyrolytic carbon surface revealed that the charged hydrophilic surface provides the optimal surface to facilitate self-assembly of SELPs.

Mechanistic understanding of monovalent cation transport in eumelanin pigments.

Recent research advances in charge-conducting materials have enabled the transformation of the naturally-occurring materials into crucial components in many technologies, including renewable energy storage devices or bioelectronics. Among various candidates, eumelanins are promising charge storage materials, exhibiting hybrid electronic ionic conductivity in a hydrated environment. The chemical and electrochemical properties of eumelanins are relatively well studied; however, the structure-property relationship is still elusive up to date. Herein, we reported the mesoscale structure of eumelanins and its impact on the charge transport. X-ray scattering suggests that eumelanin pigments exhibit the semi-crystalline structure with ordered d-spacings. These unique mesoscale structures further influence the charge transport mechanism with the cations of various sizes. Understanding the structures with consequent electrochemical properties suggest that eumelanins can further be tuned to serve as high-performance naturally-occurring charge storage materials.

Selection of DNA Cleavage Sites by Topoisomerase II Results from Enzyme-Induced Flexibility of DNA.

Topoisomerase II cleaves DNA at preferred sequences with different efficiencies; however, the mechanism of cleavage site selection is not known. Here we used single-molecule fluorescence assays that monitor several critical steps of DNA-topoisomerase II interactions, including binding/dissociation, bending/straightening, and cleavage/religation, and reveal that the cleavage site is selected mainly during the bending step. Furthermore, despite the sensitivity of the bending rate to the DNA sequence, it is not an intrinsic property of the DNA itself. Rather, it is determined by protein-DNA interactions.

Energetic Costs, Precision, and Transport Efficiency of Molecular Motors.

An efficient molecular motor would deliver cargo to the target site at a high speed and in a punctual manner while consuming a minimal amount of energy. According to a recently formulated thermodynamic principle, referred to as the thermodynamic uncertainty relation, the travel distance of a motor and its variance are, however, constrained by the free energy being consumed. Here we use the principle underlying the uncertainty relation to quantify the transport efficiency of molecular motors for varying ATP concentration ([ATP]) and applied load (f). Our analyses of experimental data find that transport efficiencies of the motors studied here are semioptimized under the cellular condition. The efficiency is significantly deteriorated for a kinesin-1 mutant that has a longer neck-linker, which underscores the importance of molecular structure. It is remarkable to recognize that, among many possible directions for optimization, biological motors have evolved to optimize the transport efficiency in particular.

Spray-Processed Composites with High Conductivity and Elasticity.

Highly conductive elastic composites were constructed using multistep solution-based fabrication methods that included the deposition of a nonwoven polymer fiber mat through solution blow spinning and nanoparticle nucleation. High nanoparticle loading was achieved by introducing silver nanoparticles into the fiber spinning solution. The presence of the silver nanoparticles facilitates improved uptake of silver nanoparticle precursor in subsequent processing steps. The precursor is used to generate a second nanoparticle population, leading to high loading and conductivity. Establishing high nanoparticle loading in a microfibrous block copolymer network generated deformable composites that can sustain electrical conductivities reaching 9000 S/cm under 100% tensile strain. These conductive elastic fabrics can retain at least 70% of their initial electrical conductivity after being stretched to 100% strain and released for 500 cycles. This composite material system has the potential to be implemented in wearable electronics and robotic systems.

Dynamic coordination of two-metal-ions orchestrates λ-exonuclease catalysis.

Metal ions at the active site of an enzyme act as cofactors, and their dynamic fluctuations can potentially influence enzyme activity. Here, we use λ-exonuclease as a model enzyme with two Mg2+ binding sites and probe activity at various concentrations of magnesium by single-molecule-FRET. We find that while MgA2+ and MgB2+ have similar binding constants, the dissociation rate of MgA2+ is two order of magnitude lower than that of MgB2+ due to a kinetic-barrier-difference. At physiological Mg2+ concentration, the MgB2+ ion near the 5'-terminal side of the scissile phosphate dissociates each-round of degradation, facilitating a series of DNA cleavages via fast product-release concomitant with enzyme-translocation. At a low magnesium concentration, occasional dissociation and slow re-coordination of MgA2+ result in pauses during processive degradation. Our study highlights the importance of metal-ion-coordination dynamics in correlation with the enzymatic reaction-steps, and offers insights into the origin of dynamic heterogeneity in enzymatic catalysis.

Physical insight into the thermodynamic uncertainty relation using Brownian motion in tilted periodic potentials.

Using Brownian motion in periodic potentials V(x) tilted by a force f, we provide physical insight into the thermodynamic uncertainty relation, a recently conjectured principle for statistical errors and irreversible heat dissipation in nonequilibrium steady states. According to the relation, nonequilibrium output generated from dissipative processes necessarily incurs an energetic cost or heat dissipation q, and in order to limit the output fluctuation within a relative uncertainty ε, at least 2k_{B}T/ε^{2} of heat must be dissipated. Our model shows that this bound is attained not only at near-equilibrium [f≪V^{'}(x)] but also at far-from-equilibrium [f≫V^{'}(x)], more generally when the dissipated heat is normally distributed. Furthermore, the energetic cost is maximized near the critical force when the barrier separating the potential wells is about to vanish and the fluctuation of Brownian particles is maximized. These findings indicate that the deviation of heat distribution from Gaussianity gives rise to the inequality of the uncertainty relation, further clarifying the meaning of the uncertainty relation. Our derivation of the uncertainty relation also recognizes a bound of nonequilibrium fluctuations that the variance of dissipated heat (σ_{q}^{2}) increases with its mean (µ_{q}), and it cannot be smaller than 2k_{B}Tµ_{q}.

Quantifying the Heat Dissipation from a Molecular Motor's Transport Properties in Nonequilibrium Steady States.

Theoretical analysis, which maps single-molecule time trajectories of a molecular motor onto unicyclic Markov processes, allows us to evaluate the heat dissipated from the motor and to elucidate its dependence on the mean velocity and diffusivity. Unlike passive Brownian particles in equilibrium, the velocity and diffusion constant of molecular motors are closely inter-related. In particular, our study makes it clear that the increase of diffusivity with the heat production is a natural outcome of active particles, which is reminiscent of the recent experimental premise that the diffusion of an exothermic enzyme is enhanced by the heat released from its own catalytic turnover. Compared with freely diffusing exothermic enzymes, kinesin-1, whose dynamics is confined on one-dimensional tracks, is highly efficient in transforming conformational fluctuations into a locally directed motion, thus displaying a significantly higher enhancement in diffusivity with its turnover rate. Putting molecular motors and freely diffusing enzymes on an equal footing, our study offers a thermodynamic basis to understand the heat-enhanced self-diffusion of exothermic enzymes.