Designing an enzyme-based nanobiosensor using molecular modeling techniques.

Nanobiosensors can be built via functionalization of atomic force microscopy (AFM) tips with biomolecules capable of interacting with the analyte on a substrate, and the detection being performed by measuring the force between the immobilized biomolecule and the analyte. The optimization of such sensors may require multiple experiments to determine suitable experimental conditions for the immobilization and detection. In this study we employ molecular modeling techniques to assist in the design of nanobiosensors to detect herbicides. As a proof of principle, the properties of acetyl co-enzyme A carboxylase (ACC) were obtained with molecular dynamics simulations, from which the dimeric form in an aqueous solution was found to be more suitable for immobilization owing to a smaller structural fluctuation than the monomeric form. Upon solving the nonlinear Poisson-Boltzmann equation using a finite-difference procedure, we found that the active sites of ACC exhibited a positive surface potential while the remainder of the ACC surface was negatively charged. Therefore, optimized biosensors should be prepared with electrostatic adsorption of ACC onto an AFM tip functionalized with positively charged groups, leaving the active sites exposed to the analyte. The preferential orientation for the herbicides diclofop and atrazine with the ACC active site was determined by molecular docking calculations which displayed an inhibition coefficient of 0.168 µM for diclofop, and 44.11 µM for atrazine. This binding selectivity for the herbicide family of diclofop was confirmed by semiempirical PM6 quantum chemical calculations which revealed that ACC interacts more strongly with the herbicide diclofop than with atrazine, showing binding energies of -119.04 and +8.40 kcal mol(-1), respectively. The initial measurements of the proposed nanobiosensor validated the theoretical calculations and displayed high selectivity for the family of the diclofop herbicides.

Antimony-doped tin oxide nanocrystals: synthesis and solubility behavior in organic solvents.

This work focuses on the nonaqueous synthesis of antimony-doped tin oxide nanocrystals in the size range of 2-6 nm and the investigation of their solubility in organic solvents (CHCl(3) and THF) in the presence of amphiphilic molecules (oleic acid and oleylamine). To unravel the underlying processes, a set of molecular dynamics simulations is performed involving the compatibility of oleic acid and oleylamine in mixtures with both CHCl(3) and THF. The results show that the method is useful for obtaining the desired oxide, and that the interaction between amphiphilic molecules and solvents can be predicted by molecular dynamics simulations with very good qualitative agreement.

Immobilization and unbinding investigation of the antigen-antibody complex using theoretical and experimental techniques.

Experimental results for the antibody known as immunoglobulin G - IgG interacting with phenobarbital were obtained via atomic force microscopy (AFM) and thereafter investigated using computer simulation modeling tools. Using molecular dynamics simulation and docking calculations, the energetically stable configurations of an immobilized antibody over a silicon surface were searched. Six stable configurations of the immobilized antibody over the silicon nitride surface covered by linker molecules were found. Although, only three of them (P1, P2, P5) maintained the Fragment antigen binding available for antigen interaction. Therefore, these configurations were equilibrated after reaching 100 ns molecular dynamics trajectory. The average interaction energy between the surface and the immunoglobulin G - IgG antibody in the P1, P2 and P5 configurations were -62.4 ±â€¯2.4 kcal/mol; -54.3 ±â€¯5.7 kcal/mol, and -360.9 ±â€¯4.2 kcal/mol respectively. Phenobarbital was docked within the Fab domain of P1, P2, and P5 immobilized configurations and equilibrated with molecular dynamics for binding energy estimation. Then, steered molecular dynamics was performed to evaluate unbinding energy pathway between phenobarbital and IgG in each of the three-oriented IgG configurations. No significant differences were observed in the rupture force values (EP1 = 591 ±â€¯13 pN, EP2 = 605 ±â€¯18 pN, and EP5 = 610 ±â€¯45 pN). In comparison, the average AFM experimental results were (641.6 ±â€¯363.3 pN). Therefore, it is worth noting that P5 is the configuration with highest protein-surface interaction. Therefore, the force value calculated for the P5 orientation is statistically more favorable and it is the one to be compared to the experimental data. The agreement between experimental and theoretical results indicates a favorable presented for this study opening new perspectives for antigen-antibody evaluation.

Design of an Internal Model Control strategy for single-phase grid-connected PWM inverters and its performance analysis with a non-linear local load and weak grid.

This paper presents the design of a controller based on Internal Model Control (IMC) applied to a grid-connected single-phase PWM inverter. The mathematical modeling of the inverter and the LCL output filter, used to project the 1-DOF IMC controller, is presented and the decoupling of grid voltage by a Feedforward strategy is analyzed. A Proportional - Resonant Controller (P+Res) was used for the control of the same plant in the running of experimental results, thus moving towards the discussion of differences regarding IMC and P+Res performances, which arrived at the evaluation of the proposed control strategy. The results are presented for typical conditions, for weak-grid and for non-linear local load, in order to verify the behavior of the controller against such situations.

Modeling the coverage of an AFM tip by enzymes and its application in nanobiosensors.

A stochastic simulation of adsorption processes was developed to simulate the coverage of an atomic force microscope (AFM) tip with enzymes represented as rigid polyhedrons. From geometric considerations of the enzyme structure and AFM tip, we could estimate the average number of active sites available to interact with substrate molecules in the bulk. The procedure was exploited to determine the interaction force between acetyl-CoA carboxylase enzyme (ACC enzyme) and its substrate diclofop, for which steered molecular dynamics (SMD) was used. The theoretical force of (1.6±0.5) nN per enzyme led to a total force in remarkable agreement with the experimentally measured force with AFM, thus demonstrating the usefulness of the procedure proposed here to assist in the interpretation of nanobiosensors experiments.

Molecular modeling of enzyme attachment on AFM probes.

The immobilization of enzymes on atomic force microscope tip (AFM tip) surface is a crucial step in the development of nanobiosensors to be used in detection process. In this work, an atomistic modeling of the attachment of the acetyl coenzyme A carboxylase (ACC enzyme) on a functionalized AFM tip surface is proposed. Using electrostatic considerations, suitable enzyme-surface orientations with the active sites of the ACC enzyme available for interactions with bulk molecules were found. A 50 ns molecular dynamics trajectory in aqueous solution was obtained and surface contact area, hydrogen bonding and protein stability were analyzed. The enzyme-surface model proposed here with minor adjustment can be applied to study antigen-antibody interactions as well as enzyme immobilization on silica for chromatography applications.

Chitosan molecular structure as a function of N-acetylation.

Molecular dynamics simulations have been carried out to characterize the structure and solubility of chitosan nanoparticle-like structures as a function of the deacetylation level (0, 40, 60, and 100%) and the spatial distribution of the N-acetyl groups in the particles. The polysaccharide chains of highly N-deacetylated particles where the N-acetyl groups are uniformly distributed present a high flexibility and preference for the relaxed two-fold helix and five-fold helix motifs. When these groups are confined to a given region of the particle, the chains adopt preferentially a two-fold helix with ϕ and ψ values close to crystalline chitin. Nanoparticles with up to 40% acetylation are moderately soluble, forming stable aggregates when the N-acetyl groups are unevenly distributed. Systems with 60% or higher N-acetylation levels are insoluble and present similar degrees of swelling regardless the distribution of their N-acetyl groups. Overall particle solvation is highly affected by electrostatic forces resulting from the degree of acetylation. The water mobility and orientation around the polysaccharide chains affects the stability of the intramolecular O3-HO3((n)) ···O5((n +) (1)) hydrogen bond, which in turn controls particle aggregation.

Interaction between the CBM of Cel9A from Thermobifida fusca and cellulose fibers.

Molecular docking and molecular dynamics (MD) simulations were used to investigate the binding of a cellodextrin chain in a crystal-like conformation to the carbohydrate-binding module (CBM) of Cel9A from Thermobifida fusca. The fiber was found to bind to the CBM in a single and well-defined configuration in-line with the catalytic cleft, supporting the hypothesis that this CBM plays a role in the catalysis by feeding the catalytic domain (CD) with a polysaccharide chain. The results also expand the current known list of residues involved in the binding. The polysaccharide-protein attachment is shown to be mediated by five amine/amide-containing residues. E478 and E559 were found not to interact directly with the sugar chain; instead they seem to be responsible to stabilize the binding motif via hydrogen bonds.

Characterization of Chitin and Chitosan Molecular Structure in Aqueous Solution.

Molecular dynamics simulations have been used to characterize the structure of single chitin and chitosan chains in aqueous solutions. Chitin chains, whether isolated or in the form of a ß-chitin nanoparticle, adopt the 2-fold helix with ϕ and φ values similar to its crystalline state. In solution, the intramolecular hydrogen bond HO3(n)···O5(n+1) responsible for the 2-fold helical motif in these polysaccharides is stabilized by hydrogen bonds with water molecules in a well-defined orientation. On the other hand, chitosan can adopt five distinct helical motifs, and its conformational equilibrium is highly dependent on pH. The hydrogen bond pattern and solvation around the O3 atom of insoluble chitosan (basic pH) are nearly identical to these quantities in chitin. Our findings suggest that the solubility and conformation of these polysaccharides are related to the stability of the intrachain HO3(n)···O5(n+1) hydrogen bond, which is affected by the water exchange around the O3-HO3 hydroxyl group.

On the relative stabilities of the alkali cations 222 cryptates in the gas phase and in water-methanol solution.

The relative stabilities of the alkali [M subset 222]+ cryptates (M = Na, K, Rb and Cs) in the gas phase and in solution (80:20 v/v methanol:water mixture) at 298 K, are computed using a combination of ab initio quantum-chemical calculations (HF/6-31G and MP2/6-31+G*//HF/6-31+G*) and explicit-solvent Monte Carlo free-energy simulations. The results suggest that the relative stabilities of the cryptates in solution are due to a combination of steric effects (compression of large ions within the cryptand cavity), electronic effects (delocalization of the ionic charge onto the cryptand atoms) and solvent effects (dominantly the ionic dessolvation penalty). Thus, the relative stabilities in solution cannot be rationalized solely on the basis of a simple match or mismatch between the ionic radius and the cryptand cavity size as has been suggested previously. For example, although the [K subset 222]+ cryptate is found to be the most stable in solution, in agreement with experimental data, it is the [Na subset 222]+ cryptate that is the most stable in the gas phase. The present results provide further support to the notion that the solvent in which supramolecules are dissolved plays a key role in modulating molecular recognition processes.