Electrochemical Energy Harvesting Using Microbial Active Matter.

With the continuous growth in world population and economy, the global energy demand is increasing rapidly. Given that non-renewable energy sources will eventually deplete, there is increasing need for clean, alternative renewable energy sources, which will be inexpensive and involve minimum risk of environmental pollution. In this paper, harnessing the activity of cupric reductase NDH-2 enzyme present in Escherichia coli bacterial cells, we demonstrate a simple and efficient energy harvesting strategy within an electrochemical chamber without the requirement of any external fuels or force fields. The transduction of energy has been demonstrated with various strains of E. coli, indicating that this strategy could, in principle, be applicable for other microbial catalytic systems. We offer a simple mechanism of the energy transduction process considering the bacterial enzyme-mediated redox reaction occurring over the working electrode of the electrochemical cell. Also, the amount of energy generated has been found to be depending on the motility of bacteria within the experimental chamber, suggesting possible opportunities for developing microbial motility-controlled small scale power generators. Finally, we show that the Faradaic electrochemical energy harvested is large enough to power a commercial light emitting diode connected to an amplifier circuit. We expect the present study to generate sufficient interest within soft condensed matter and biophysics communities, and offer useful platforms for controlled energy generation at the small scales.

Micro-Bio-Chemo-Mechanical-Systems: Micromotors, Microfluidics, and Nanozymes for Biomedical Applications.

Wireless nano-/micromotors powered by chemical reactions and/or external fields generate motive forces, perform tasks, and significantly extend short-range dynamic responses of passive biomedical microcarriers. However, before micromotors can be translated into clinical use, several major problems, including the biocompatibility of materials, the toxicity of chemical fuels, and deep tissue imaging methods, must be solved. Nanomaterials with enzyme-like characteristics (e.g., catalase, oxidase, peroxidase, superoxide dismutase), that is, nanozymes, can significantly expand the scope of micromotors' chemical fuels. A convergence of nanozymes, micromotors, and microfluidics can lead to a paradigm shift in the fabrication of multifunctional micromotors in reasonable quantities, encapsulation of desired subsystems, and engineering of FDA-approved core-shell structures with tuneable biological, physical, chemical, and mechanical properties. Microfluidic methods are used to prepare stable bubbles/microbubbles and capsules integrating ultrasound, optoacoustic, fluorescent, and magnetic resonance imaging modalities. The aim here is to discuss an interdisciplinary approach of three independent emerging topics: micromotors, nanozymes, and microfluidics to creatively: 1) embrace new ideas, 2) think across boundaries, and 3) solve problems whose solutions are beyond the scope of a single discipline toward the development of micro-bio-chemo-mechanical-systems for diverse bioapplications.

Enhanced Diffusion of Passive Tracers in Active Enzyme Solutions.

Colloidal suspensions containing microscopic swimmers have been the focus of recent studies aimed at understanding the principles of energy transfer in fluidic media at low Reynolds number conditions. Going down in scale, active enzymes have been shown to be force-generating, nonequilibrium systems, thus offering opportunity to examine energy transfer at the ultralow Reynolds number regime. By monitoring the change of diffusion of inert tracers dispersed in active enzyme solutions, we demonstrate that the nature of energy transfer in these systems is similar to that reported for larger microscopic active systems, despite the large differences in scale, modes of energy transduction, and propulsion. Additionally, even an enzyme that catalyzes an endothermic reaction behaves analogously, suggesting that heat generation is not the primary factor for the observed enhanced tracer diffusion. Our results provide new insights into the mechanism of energy transfer at the molecular level.

Exothermicity Is Not a Necessary Condition for Enhanced Diffusion of Enzymes.

Recent experiments have revealed that the diffusivity of exothermic and fast enzymes is enhanced when they are catalytically active, and different physical mechanisms have been explored and quantified to account for this observation. We perform measurements on the endothermic and relatively slow enzyme aldolase, which also shows substrate-induced enhanced diffusion. We propose a new physical paradigm, which reveals that the diffusion coefficient of a model enzyme hydrodynamically coupled to its environment increases significantly when undergoing changes in conformational fluctuations in a substrate concentration dependent manner, and is independent of the overall turnover rate of the underlying enzymatic reaction. Our results show that substrate-induced enhanced diffusion of enzyme molecules can be explained within an equilibrium picture and that the exothermicity of the catalyzed reaction is not a necessary condition for the observation of this phenomenon.

Micromotors Powered by Enzyme Catalysis.

Active biocompatible systems are of great current interest for their possible applications in drug or antidote delivery at specific locations. Herein, we report the synthesis and study of self-propelled microparticles powered by enzymatic reactions and their directed movement in substrate concentration gradient. Polystyrene microparticles were functionalized with the enzymes urease and catalase using a biotin-streptavidin linkage procedure. The motion of the enzyme-coated particles was studied in the presence of the respective substrates, using optical microscopy and dynamic light scattering analysis. The diffusion of the particles was found to increase in a substrate concentration dependent manner. The directed chemotactic movement of these enzyme-powered motors up the substrate gradient was studied using three-inlet microfluidic channel architecture.

Impulsive Enzymes: A New Force in Mechanobiology.

We review studies that quantify newly discovered forces from single enzymatic reactions. These forces arise from the conversion of chemical energy to kinetic energy, which can be harnessed to direct diffusion of the enzyme up a concentration gradient of substrate, a novel phenomenon of molecular chemotaxis. When immobilized, enzymes can move fluid around them and perform directional pumping in microfluidic chambers. Because of the extensive array of enzymes in biological cells, we also develop three new hypotheses: that enzymatic self diffusion can assist in organizing signaling pathways in cells, can assist in pumping of fluid in cells, and can impose biologically significant forces on organelles, which will be manifested as stochastic motion not explained by thermal forces or myosin II. Such mechanochemical phenomena open up new directions in research in mechanobiology in which all enzymes, in addition to their primary function as catalysts for reactions, may have secondary functions as initiators of mechanosensitive transduction pathways.

Two-dimensional NMR measurement and point dipole model prediction of paramagnetic shift tensors in solids.

A new two-dimensional Nuclear Magnetic Resonance (NMR) experiment to separate and correlate the first-order quadrupolar and chemical/paramagnetic shift interactions is described. This experiment, which we call the shifting-d echo experiment, allows a more precise determination of tensor principal components values and their relative orientation. It is designed using the recently introduced symmetry pathway concept. A comparison of the shifting-d experiment with earlier proposed methods is presented and experimentally illustrated in the case of (2)H (I = 1) paramagnetic shift and quadrupolar tensors of CuCl2⋅2D2O. The benefits of the shifting-d echo experiment over other methods are a factor of two improvement in sensitivity and the suppression of major artifacts. From the 2D lineshape analysis of the shifting-d spectrum, the (2)H quadrupolar coupling parameters are 〈Cq〉 = 118.1 kHz and 〈ηq〉 = 0.88, and the (2)H paramagnetic shift tensor anisotropy parameters are 〈ζP〉 = - 152.5 ppm and 〈ηP〉 = 0.91. The orientation of the quadrupolar coupling principal axis system (PAS) relative to the paramagnetic shift anisotropy principal axis system is given by (α,ß,γ)=(π2,π2,0). Using a simple ligand hopping model, the tensor parameters in the absence of exchange are estimated. On the basis of this analysis, the instantaneous principal components and orientation of the quadrupolar coupling are found to be in excellent agreement with previous measurements. A new point dipole model for predicting the paramagnetic shift tensor is proposed yielding significantly better agreement than previously used models. In the new model, the dipoles are displaced from nuclei at positions associated with high electron density in the singly occupied molecular orbital predicted from ligand field theory.

Self-powered enzyme micropumps.

Non-mechanical nano- and microscale pumps that function without the aid of an external power source and provide precise control over the flow rate in response to specific signals are needed for the development of new autonomous nano- and microscale systems. Here we show that surface-immobilized enzymes that are independent of adenosine triphosphate function as self-powered micropumps in the presence of their respective substrates. In the four cases studied (catalase, lipase, urease and glucose oxidase), the flow is driven by a gradient in fluid density generated by the enzymatic reaction. The pumping velocity increases with increasing substrate concentration and reaction rate. These rechargeable pumps can be triggered by the presence of specific analytes, which enables the design of enzyme-based devices that act both as sensor and pump. Finally, we show proof-of-concept enzyme-powered devices that autonomously deliver small molecules and proteins in response to specific chemical stimuli, including the release of insulin in response to glucose.

DNA polymerase as a molecular motor and pump.

DNA polymerase is responsible for synthesizing DNA, a key component in the running of biological machinery. Using fluorescence correlation spectroscopy, we demonstrate that the diffusive movement of a molecular complex of DNA template and DNA polymerase enhances during nucleotide incorporation into the growing DNA template. The diffusion coefficient of the complex also shows a strong dependence on its inorganic cofactor, Mg2+ ions. When exposed to gradients of either nucleotide or cofactor concentrations, an ensemble of DNA polymerase complex molecules shows collective movement toward regions of higher concentrations. By immobilizing the molecular complex on a patterned gold surface, we demonstrate the fabrication of DNA polymerase-powered fluid pumps. These miniature pumps are capable of transporting fluid and tracer particles in a directional manner with the pumping speed increasing in the presence of the cofactor. The role of DNA polymerase as a micropump opens up avenues for designing miniature fluid pumps using enzymes as engines.

Investigations into variations in local cationic environment in layered oxide series InGaO3(ZnO)m (m = 1-4).

Layered oxides of the series InGaO3(ZnO)m (m = 1-4) are interesting due to their structural anisotropy. Here, we report a comprehensive study of their structural details, focusing on the local cationic environment in bulk powder samples by MASNMR and EXAFS, which is hitherto not attempted. It is found that the Ga geometry varies gradually from pure pentacoordinated to a mixture of penta and tetracoordinated with increasing amounts of tetracoordination as we move across the series, contrary to previous reports suggesting exclusively trigonal bipyramidal coordination in all the compounds. A similar observation is also made in the case of Zn and structural evolution involving the dissolution of Ga in a ZnO4 tetrahedral network in a sandwich layer can be discerned, as the insulating ZnO layer size increases.

Sideband separation experiments in NMR with phase incremented echo train acquisition.

A general approach for enhancing sensitivity of nuclear magnetic resonance sideband separation experiments, such as Two-Dimensional One Pulse (TOP), Magic-Angle Turning (MAT), and Phase Adjust Spinning Sidebands (PASS) experiments, with phase incremented echo-train acquisition (PIETA) is described. This approach is applicable whenever strong inhomogeneous broadenings dominate the unmodulated frequency resonances, such as in non-crystalline solids or in samples with large residual frequency anisotropy. PIETA provides significant sensitivity enhancements while also eliminating spectral artifacts would normally be present with Carr-Purcell-Meiboom-Gill acquisition. Additionally, an intuitive approach is presented for designing and processing echo train acquisition magnetic resonance experiments on rotating samples. Affine transformations are used to relate the two-dimensional signals acquired in TOP, MAT, and PASS experiments to a common coordinate system. Depending on sequence design and acquisition conditions two significant artifacts can arise from truncated acquisition time and discontinuous damping in the T2 decay. Here we show that the former artifact can always be eliminated through selection of a suitable affine transformation, and give the conditions in which the latter can be minimized or removed entirely.

Enzyme molecules as nanomotors.

Using fluorescence correlation spectroscopy, we show that the diffusive movements of catalase enzyme molecules increase in the presence of the substrate, hydrogen peroxide, in a concentration-dependent manner. Employing a microfluidic device to generate a substrate concentration gradient, we show that both catalase and urease enzyme molecules spread toward areas of higher substrate concentration, a form of chemotaxis at the molecular scale. Using glucose oxidase and glucose to generate a hydrogen peroxide gradient, we induce the migration of catalase toward glucose oxidase, thereby showing that chemically interconnected enzymes can be drawn together.

A catalytically driven organometallic molecular motor.

We have observed by NMR spectroscopy that the diffusive movement of a ruthenium-based Grubbs' catalyst increases during ring-closing metathesis as a function of the substrate concentration. This is one of the smallest single molecule motors to exhibit catalytically driven motion.

Trading sensitivity for information: Carr-Purcell-Meiboom-Gill acquisition in solid-state NMR.

The Carr-Purcell-Meiboom-Gill (CPMG) experiment has gained popularity in solid-state NMR as a method for enhancing sensitivity for anisotropically broadened spectra of both spin 1/2 and half integer quadrupolar nuclei. Most commonly, the train of CPMG echoes is Fourier transformed directly, which causes the NMR powder pattern to break up into a series of sidebands, sometimes called "spikelets." Larger sensitivity enhancements are observed as the delay between the pi pulses is shortened. As the duration between the pi pulses is shortened, however, the echoes become truncated and information about the nuclear spin interactions is lost. We explored the relationship between enhanced sensitivity and loss of information as a function of the product Omega 2tau, where Omega is the span of the anisotropic lineshape and 2tau is the pi pulse spacing. For a lineshape dominated by the nuclear shielding anisotropy, we found that the minimum uncertainty in the tensor values is obtained using Omega 2tau values in the range Omega 2tau approximately 12(-1)(+6) and Omega 2tau approximately 9(-3)(+3) for eta(s)=0 and eta(s)=1, respectively. For an anisotropic second-order quadrupolar central transition lineshape under magic-angle spinning (MAS), the optimum range of Omega 2tau approximately 9(-2)(+3) was found. Additionally, we show how the Two-dimensional One Pulse (TOP) like processing approach can be used to eliminate the cumbersome sideband pattern lineshape and recover a more familiar lineshape that is easily analyzed with conventional lineshape simulation algorithms.

Optimum excitation of "enhanced" central transition populations.

Central transition (CT) sensitivity enhancement schemes that transfer polarization from satellites to the CT through selective saturation or inversion of neighboring satellite transitions have provided a welcome improvement for magic-angle spinning spectra of half-integer quadrupole nuclei. While many researchers have investigated and developed different methods of creating enhanced CT populations, here we investigate the conversion of these enhanced CT populations into observable CT coherence. We show a somewhat unexpected result that a conversion pulse length optimized for maximum sensitivity on equilibrium populations may not be optimum for an enhanced (non-equilibrium) polarization. Furthermore, CT enhancements can be lost if excessive rf field strength is used to convert this enhanced polarization into CT coherence. While a maximally enhanced CT signal is expected when using a perfectly selective CT conversion pulse, we have found that significant sensitivity loss can occur when using surprisingly low rf field strengths, even for sites with relatively large quadrupole coupling constants. We have systematically investigated these issues, and present some general guidelines and expectations when optimizing the conversion of enhanced (non-equilibrium) CT populations into observable CT coherence.

Spectral editing in solid-state MAS NMR of quadrupolar nuclei using selective satellite inversion.

A sensitivity enhancement method based on selective adiabatic inversion of a satellite transition has been employed in a (pi/2)CT-(pi)ST1-(pi/2)CT spectral editing sequence to both enhance and resolve multisite NMR spectra of quadrupolar nuclei. In addition to a total enhancement of 2.5 times for spin 3/2 nuclei, enhancements up to 2.0 times is reported for the edited sites in a mixture of rubidium salts.