Regioselectivity mechanism of the Thunbergia alata Δ6-16:0-acyl carrier protein desaturase.

Plant plastidial acyl-acyl carrier protein (ACP) desaturases are a soluble class of diiron-containing enzymes that are distinct from the diiron-containing integral membrane desaturases found in plants and other organisms. The archetype of this class is the stearoyl-ACP desaturase which converts stearoyl-ACP into oleoyl (18:1Δ9cis)-ACP. Several variants expressing distinct regioselectivity have been described including a Δ6-16:0-ACP desaturase from black-eyed Susan vine (Thunbergia alata). We solved a crystal structure of the T. alata desaturase at 2.05 Å resolution. Using molecular dynamics (MD) simulations, we identified a low-energy complex between 16:0-ACP and the desaturase that would position C6 and C7 of the acyl chain adjacent to the diiron active site. The model complex was used to identify mutant variants that could convert the T. alata Δ6 desaturase to Δ9 regioselectivity. Additional modeling between ACP and the mutant variants confirmed the predicted regioselectivity. To validate the in-silico predictions, we synthesized two variants of the T. alata desaturase and analyzed their reaction products using gas chromatography-coupled mass spectrometry. Assay results confirmed that mutants designed to convert T. alata Δ6 to Δ9 selectivity exhibited the predicted changes. In complementary experiments, variants of the castor desaturase designed to convert Δ9 to Δ6 selectivity lost some of their Δ9 desaturation ability and gained the ability to desaturate at the Δ6 position. The computational workflow for revealing the mechanistic understanding of regioselectivity presented herein lays a foundation for designing acyl-ACP desaturases with novel selectivities to increase the diversity of monoenes available for bioproduct applications.

Protonation of Serine in Gas and Condensed and Microsolvated States in Aqueous Solution.

Identification of molecules and elucidation of their chemical structure are ubiquitous problems in chemistry. Mass spectrometry (MS) can be used due to its sensitivity and versatility. For detection to occur, analytes must be ionized and transferred to the gas phase. Soft ionization processes such as electrospray ionization are popular; however, resulting microsolvated phases can alter the chemistry of analytes and therefore detection and identification. To understand these processes, we use computational methods to probe the ionization propensity of serine in the gas phase, aqueous microsolvated clusters, and aqueous solution. We show that the tautomeric form of serine is altered by the presence of water, as five water molecules can stabilize the zwitterionic tautomer. Inclusion of cosolutes such as ions can stabilize the zwitterion with as few as one or two water molecules present. We demonstrate that ionization propensity, as measured by gas phase bacisity, can increase by over 100 kJ/mol when placed in a small water-serine cluster, showing the sensitivity of the chemistry of microsolvated analytes. Finally, detailed analysis reveals that small droplets (less than seven water molecules) are extremely sensitive to addition of further water molecules. Beyond this limit, structural and electronic properties change little with droplet size.

Molecular-level origin of the carboxylate head group response to divalent metal ion complexation at the air-water interface.

We exploit gas-phase cluster ion techniques to provide insight into the local interactions underlying divalent metal ion-driven changes in the spectra of carboxylic acids at the air-water interface. This information clarifies the experimental findings that the CO stretching bands of long-chain acids appear at very similar energies when the head group is deprotonated by high subphase pH or exposed to relatively high concentrations of Ca2+ metal ions. To this end, we report the evolution of the vibrational spectra of size-selected [Ca2+·RCO2-]+·(H2O) n=0to12 and RCO2-·(H2O) n=0to14 cluster ions toward the features observed at the air-water interface. Surprisingly, not only does stepwise hydration of the RCO2- anion and the [Ca2+·RCO2-]+ contact ion pair yield solvatochromic responses in opposite directions, but in both cases, the responses of the 2 (symmetric and asymmetric stretching) CO bands to hydration are opposite to each other. The result is that both CO bands evolve toward their interfacial asymptotes from opposite directions. Simulations of the [Ca2+·RCO2-]+·(H2O) n clusters indicate that the metal ion remains directly bound to the head group in a contact ion pair motif as the asymmetric CO stretch converges at the interfacial value by n = 12. This establishes that direct metal complexation or deprotonation can account for the interfacial behavior. We discuss these effects in the context of a model that invokes the water network-dependent local electric field along the C-C bond that connects the head group to the hydrocarbon tail as the key microscopic parameter that is correlated with the observed trends.

Atomistic insight on structure and dynamics of spinach acyl carrier protein with substrate length.

The plant acyl-acyl carrier protein (ACP) desaturases are a family of soluble enzymes that convert saturated fatty acyl-ACPs into their cis-monounsaturated equivalents in an oxygen-dependent reaction. These enzymes play a key role in biosynthesis of monounsaturated fatty acids in plants. ACPs are central proteins in fatty acid biosynthesis that deliver acyl chains to desaturases. They have been reported to show a varying degree of local dynamics and structural variability depending on the acyl chain size. It has been suggested that substrate-specific changes in ACP structure and dynamics have a crucial impact on the desaturase enzymatic activity. Using molecular dynamics simulations, we investigated the intrinsic solution structure and dynamics of ACP from spinach with four different acyl chains: capric (C10), myristic (C14), palmitic (C16), and stearic (C18) acids. We found that the fatty acids can adopt two distinct structural binding motifs, which feature different binding free energies and influence the ACP dynamics in a different manner. Docking simulations of ACP to castor Δ9-desaturase and ivy Δ4-desaturase suggest that ACP desaturase interactions could lead to a preferential selection between the motifs.

Visualization of Aluminum Ions at the Mica Water Interface Links Hydrolysis State-to-Surface Potential and Particle Adhesion.

When hydrolyzable cations such as aluminum interact with solid-water interfaces, macroscopic interfacial properties (e.g., surface charge and potential) and interfacial phenomena (e.g., particle adhesion) become tightly linked with the microscopic details of ion adsorption and speciation. We use in situ atomic force microscopy to directly image individual aluminum ions at a mica-water interface and show how adsorbate populations change with pH and aluminum activity. Complementary streaming potential measurements then allow us to build a triple layer model (TLM) that links surface potentials to adsorbate populations, via equilibrium binding constants. Our model predicts that hydrolyzed species dominate the mica-water interface, even when unhydrolyzed species dominate the solution. Ab initio molecular dynamics (AIMD) simulations confirm that aluminum hydrolysis is strongly promoted at the interface. The TLM indicates that hydrolyzed adsorbates are responsible for surface-potential inversions, and we find strong correlations between hydrolyzed adsorbates and particle-adhesion forces, suggesting that these species mediate adhesion by chemical bridging.

Quantifying the hydration structure of sodium and potassium ions: taking additional steps on Jacob's Ladder.

The ability to reproduce the experimental structure of water around the sodium and potassium ions is a key test of the quality of interaction potentials due to the central importance of these ions in a wide range of important phenomena. Here, we simulate the Na+ and K+ ions in bulk water using three density functional theory functionals: (1) the generalized gradient approximation (GGA) based dispersion corrected revised Perdew, Burke, and Ernzerhof functional (revPBE-D3) (2) the recently developed strongly constrained and appropriately normed (SCAN) functional (3) the random phase approximation (RPA) functional for potassium. We compare with experimental X-ray diffraction (XRD) and X-ray absorption fine structure (EXAFS) measurements to demonstrate that SCAN accurately reproduces key structural details of the hydration structure around the sodium and potassium cations, whereas revPBE-D3 fails to do so. However, we show that SCAN provides a worse description of pure water in comparison with revPBE-D3. RPA also shows an improvement for K+, but slow convergence prevents rigorous comparison. Finally, we analyse cluster energetics to show SCAN and RPA have smaller fluctuations of the mean error of ion-water cluster binding energies compared with revPBE-D3.

Solvent reaction coordinate for an SN2 reaction.

We study the prototypical SN2 reaction Cl- + CH3Cl → CH3Cl + Cl- in water using quantum mechanics/molecular mechanics (QM/MM) computer simulations with transition path sampling and inertial likelihood maximization. We have identified a new solvent coordinate to complement the original atom-exchange coordinate used in the classic analysis by Chandrasekhar, Smith, and Jorgensen [J. Am. Chem. Soc. 107, 154 (1985)]. The new solvent coordinate quantifies instantaneous solvent-induced polarization relative to the equilibrium average charge density at each point along the reaction pathway. On the basis of likelihood scores and committor distributions, the new solvent coordinate improves upon the description of solvent dynamical effects relative to previously proposed solvent coordinates. However, it does not increase the transmission coefficient or the accuracy of a transition state theory rate calculation.

Direct Observation of the Orientational Anisotropy of Buried Hydroxyl Groups inside Muscovite Mica.

Muscovite mica (001) is a widely used model surface for controlling molecular assembly and a common substrate for environmental adsorption processes. The mica (001) surface displays near-trigonal symmetry, but many molecular adsorbates-including water-exhibit unequal probabilities of alignment along its three nominally equivalent lattice directions. Buried hydroxyl groups within the muscovite structure are speculated to be responsible, but direct evidence is lacking. Here, we utilize vibrational sum frequency generation spectroscopy (vSFG) to characterize the orientation and hydrogen-bonding environment of near-surface hydroxyls inside mica. Multiple distinct peaks are detected in the O-H stretch region, which we attribute to Si/Al substitution in the SiO4 tetrahedron and K+ ion adsorption above the hydroxyls based on density functional theory simulations. Our findings demonstrate that vSFG can identify the absolute orientation of -OH groups and, hence, the surface termination at a mica surface, providing a means to investigate how -OH groups influence molecular adsorption and better understand mica stacking-sequences and physical behavior.

Tuning crystallization pathways through sequence engineering of biomimetic polymers.

Two-step nucleation pathways in which disordered, amorphous, or dense liquid states precede the appearance of crystalline phases have been reported for a wide range of materials, but the dynamics of such pathways are poorly understood. Moreover, whether these pathways are general features of crystallizing systems or a consequence of system-specific structural details that select for direct versus two-step processes is unknown. Using atomic force microscopy to directly observe crystallization of sequence-defined polymers, we show that crystallization pathways are indeed sequence dependent. When a short hydrophobic region is added to a sequence that directly forms crystalline particles, crystallization instead follows a two-step pathway that begins with the creation of disordered clusters of 10-20 molecules and is characterized by highly non-linear crystallization kinetics in which clusters transform into ordered structures that then enter the growth phase. The results shed new light on non-classical crystallization mechanisms and have implications for the design of self-assembling polymer systems.

Peptoid Backbone Flexibilility Dictates Its Interaction with Water and Surfaces: A Molecular Dynamics Investigation.

Peptoids are peptide-mimetic biopolymers that are easy to synthesize and adaptable for use in drugs, chemical scaffolds, and coatings. However, there is insufficient information about their structural preferences and interactions with the environment in various applications. We conducted a study to understand the fundamental differences between peptides and peptoids using molecular dynamics simulations with semiempirical (PM6) and empirical (AMBER) potentials, in conjunction with metadynamics enhanced sampling. From studies of single molecules in water and on surfaces, we found that sarcosine (model peptoid) is much more flexible than alanine (model peptide) in different environments. However, the sarcosine and alanine interact similarly with a hydrophobic or a hydrophilic. Finally, this study highlights the conformational landscape of peptoids and the dominant interactions that drive peptoids toward these conformations.

Understanding the scale of the single ion free energy: A critical test of the tetra-phenyl arsonium and tetra-phenyl borate assumption.

The tetra-phenyl arsonium and tetra-phenyl borate (TATB) assumption is a commonly used extra-thermodynamic assumption that allows single ion free energies to be split into cationic and anionic contributions. The assumption is that the values for the TATB salt can be divided equally. This is justified by arguing that these large hydrophobic ions will cause a symmetric response in water. Experimental and classical simulation work has raised potential flaws with this assumption, indicating that hydrogen bonding with the phenyl ring may favor the solvation of the TB- anion. Here, we perform ab initio molecular dynamics simulations of these ions in bulk water demonstrating that there are significant structural differences. We quantify our findings by reproducing the experimentally observed vibrational shift for the TB- anion and confirm that this is associated with hydrogen bonding with the phenyl rings. Finally, we demonstrate that this results in a substantial energetic preference of the water to solvate the anion. Our results suggest that the validity of the TATB assumption, which is still widely used today, should be reconsidered experimentally in order to properly reference single ion solvation free energy, enthalpy, and entropy.

Detecting the undetectable: The role of trace surfactant in the Jones-Ray effect.

The surface tension of dilute salt water is a fundamental property that is crucial to understanding the complexity of many aqueous phase processes. Small ions are known to be repelled from the air-water surface leading to an increase in the surface tension in accordance with the Gibbs adsorption isotherm. The Jones-Ray effect refers to the observation that at extremely low salt concentration, the surface tension decreases. Determining the mechanism that is responsible for this Jones-Ray effect is important for theoretically predicting the distribution of ions near surfaces. Here we use both experimental surface tension measurements and numerical solution of the Poisson-Boltzmann equation to demonstrate that very low concentrations of surfactant in water create a Jones-Ray effect. We also demonstrate that the low concentrations of the surfactant necessary to create the Jones-Ray effect are too small to be detectable by surface sensitive spectroscopic measurements. The effect of surface curvature on this behavior is also examined, and the implications for unexplained bubble phenomena are discussed. This work suggests that the purity standards for water may be inadequate and that the interactions between ions with background impurities are important to incorporate into our understanding of the driving forces that give rise to the speciation of ions at interfaces.

Electrostatic solvation free energies of charged hard spheres using molecular dynamics with density functional theory interactions.

Determining the solvation free energies of single ions in water is one of the most fundamental problems in physical chemistry and yet many unresolved questions remain. In particular, the ability to decompose the solvation free energy into simple and intuitive contributions will have important implications for models of electrolyte solution. Here, we provide definitions of the various types of single ion solvation free energies based on different simulation protocols. We calculate solvation free energies of charged hard spheres using density functional theory interaction potentials with molecular dynamics simulation and isolate the effects of charge and cavitation, comparing to the Born (linear response) model. We show that using uncorrected Ewald summation leads to unphysical values for the single ion solvation free energy and that charging free energies for cations are approximately linear as a function of charge but that there is a small non-linearity for small anions. The charge hydration asymmetry for hard spheres, determined with quantum mechanics, is much larger than for the analogous real ions. This suggests that real ions, particularly anions, are significantly more complex than simple charged hard spheres, a commonly employed representation.

Mass density fluctuations in quantum and classical descriptions of liquid water.

First principles molecular dynamics simulation protocol is established using revised functional of Perdew-Burke-Ernzerhof (revPBE) in conjunction with Grimme's third generation of dispersion (D3) correction to describe the properties of water at ambient conditions. This study also demonstrates the consistency of the structure of water across both isobaric (NpT) and isothermal (NVT) ensembles. Going beyond the standard structural benchmarks for liquid water, we compute properties that are connected to both local structure and mass density fluctuations that are related to concepts of solvation and hydrophobicity. We directly compare our revPBE results to the Becke-Lee-Yang-Parr (BLYP) plus Grimme dispersion corrections (D2) and both the empirical fixed charged model (SPC/E) and many body interaction potential model (MB-pol) to further our understanding of how the computed properties herein depend on the form of the interaction potential.

Simulation and theory of ions at atmospherically relevant aqueous liquid-air interfaces.

Chemistry occurring at or near the surface of aqueous droplets and thin films in the atmosphere influences air quality and climate. Molecular dynamics simulations are becoming increasingly useful for gaining atomic-scale insight into the structure and reactivity of aqueous interfaces in the atmosphere. Here we review simulation studies of atmospherically relevant aqueous liquid-air interfaces, with an emphasis on ions that play important roles in the chemistry of atmospheric aerosols. In addition to surveying results from simulation studies, we discuss challenges to the refinement and experimental validation of the methodology for simulating ion adsorption to the air-water interface and recent advances in elucidating the driving forces for adsorption. We also review the recent development of a dielectric continuum theory capable of reproducing simulation and experimental data on ion behavior at aqueous interfaces.

Molecular mechanism of trehalose 6-phosphate inhibition of the plant metabolic sensor kinase SnRK1.

SUCROSE-NON-FERMENTING1-RELATED PROTEIN KINASE1 (SnRK1), a central plant metabolic sensor kinase, phosphorylates its target proteins, triggering a global shift from anabolism to catabolism. Molecular modeling revealed that upon binding of KIN10 to GEMINIVIRUS REP-INTERACTING KINASE1 (GRIK1), KIN10's activation T-loop reorients into GRIK1's active site, enabling its phosphorylation and activation. Trehalose 6-phosphate (T6P) is a proxy for cellular sugar status and a potent inhibitor of SnRK1. T6P binds to KIN10, a SnRK1 catalytic subunit, weakening its affinity for GRIK1. Here, we investigate the molecular details of T6P inhibition of KIN10. Molecular dynamics simulations and in vitro phosphorylation assays identified and validated the T6P binding site on KIN10. Under high-sugar conditions, T6P binds to KIN10, blocking the reorientation of its activation loop and preventing its phosphorylation and activation by GRIK1. Under these conditions, SnRK1 maintains only basal activity levels, minimizing phosphorylation of its target proteins, thereby facilitating a general shift from catabolism to anabolism.

Influence of Peptoid Sequence on the Mechanisms and Kinetics of 2D Assembly.

Two-dimensional (2D) materials have attracted intense interest due to their potential for applications in fields ranging from chemical sensing to catalysis, energy storage, and biomedicine. Recently, peptoids, a class of biomimetic sequence-defined polymers, have been found to self-assemble into 2D crystalline sheets that exhibit unusual properties, such as high chemical stability and the ability to self-repair. The structure of a peptoid is close to that of a peptide except that the side chains are appended to the amide nitrogen rather than the α carbon. In this study, we investigated the effect of peptoid sequence on the mechanism and kinetics of 2D assembly on mica surfaces using in situ AFM and time-resolved X-ray scattering. We explored three distinct peptoid sequences that are amphiphilic in nature with hydrophobic and hydrophilic blocks and are known to self-assemble into 2D sheets. The results show that their assembly on mica starts with deposition of aggregates that spread to establish 2D islands, which then grow by attachment of peptoids, either monomers or unresolvable small oligomers, following well-known laws of crystal step advancement. Extraction of the solubility and kinetic coefficient from the dependence of the growth rate on peptoid concentration reveals striking differences between the sequences. The sequence with the slowest growth rate in bulk and with the highest solubility shows almost no detachment; i.e., once a growth unit attaches to the island edge, there is almost no probability of detaching. Furthermore, a peptoid sequence with a hydrophobic tail conjugated to the final carboxyl residue in the hydrophilic block has enhanced hydrophobic interactions and exhibits rapid assembly both in the bulk and on mica. These assembly outcomes suggest that, while the π-π interactions between adjacent hydrophobic blocks play a major role in peptoid assembly, sequence details, particularly the location of charged groups, as well as interaction with the underlying substrate can significantly alter the thermodynamic stability and assembly kinetics.

Thermodynamics of iodide adsorption at the instantaneous air-water interface.

We performed molecular dynamics simulations using both polarizable and non-polarizable force fields to study the adsorption of iodide to the air-water interface. A novel aspect of our analysis is that the progress of ion adsorption is measured as the distance from the instantaneous interface, which is defined by a coarse-graining scheme proposed recently by Willard and Chandler ["Instantaneous liquid interfaces," J. Phys. Chem. B 114, 1954-1958 (2010)]. Referring structural and thermodynamic quantities to the instantaneous interface unmasks molecular-scale details that are obscured by thermal fluctuations when the same quantities are referred to an average measure of the position of the interface, such as the Gibbs dividing surface. Our results suggest that an ion adsorbed at the interface resides primarily in the topmost water layer, and the interfacial location of the ion is favored by enthalpy and opposed by entropy.

Development of a Systematic and Extensible Force Field for Peptoids (STEPs).

Peptoids (N-substituted glycines) are a class of biomimetic polymers that have attracted significant attention due to their accessible synthesis and enzymatic and thermal stability relative to their naturally occurring counterparts (polypeptides). While these polymers provide the promise of more robust functional materials via hierarchical approaches, they present a new challenge for computational structure prediction for material design. The reliability of calculations hinges on the accuracy of interactions represented in the force field used to model peptoids. For proteins, structure prediction based on sequence and de novo design has made dramatic progress in recent years; however, these models are not readily transferable for peptoids. Current efforts to develop and implement peptoid-specific force fields are spread out, leading to replicated efforts and a fragmented collection of parameterized sidechains. Here, we developed a peptoid-specific force field containing 70 different side chains, using GAFF2 as starting point. The new model is validated based on the generation of Ramachandran-like plots from DFT optimization compared against force field reproduced potential energy and free energy surfaces as well as the reproduction of equilibrium cis/trans values for some residues experimentally known to form helical structures. Equilibrium cis/trans distributions (Kct) are estimated for all parameterized residues to identify which residues have an intrinsic propensity for cis or trans states in the monomeric state.

Re-examining the properties of the aqueous vapor-liquid interface using dispersion corrected density functional theory.

First-principles molecular dynamics simulations, in which the forces are computed from electronic structure calculations, have great potential to provide unique insight into structure, dynamics, electronic properties, and chemistry of interfacial systems that is not available from empirical force fields. The majority of current first-principles simulations are driven by forces derived from density functional theory with generalized gradient approximations to the exchange-correlation energy, which do not capture dispersion interactions. We have carried out first-principles molecular dynamics simulations of air-water interfaces employing a particular generalized gradient approximation to the exchange-correlation functional (BLYP), with and without empirical dispersion corrections. We assess the utility of the dispersion corrections by comparison of a variety of structural, dynamic, and thermodynamic properties of bulk and interfacial water with experimental data, as well as other first-principles and force field-based simulations.