Proton network flexibility enables robustness and large electric fields in the ketosteroid isomerase active site

Hydrogen bond networks play vital roles in biological functions ranging from protein folding to enzyme catalysis. Here we combine electronic structure calculations and ab initio path integral molecular dynamics simulations, which incorporate both nuclear and electronic quantum effects, to show why the network of short hydrogen bonds in the active site of ketosteroid isomerase is remarkably robust to mutations along the network and how this gives rise to large local electric fields. We demonstrate that these properties arise from the network's ability to respond to a perturbation by shifting proton positions and redistributing electronic charge density. This flexibility leads to small changes in properties such as the partial ionization of residues and $pK_a$ isotope effects upon mutation of the residues, consistent with recent experiments. This proton flexibility is further enhanced when an extended hydrogen bond network forms in the presence of an intermediate analog, which allows us to explain the chemical origins of the large electric fields in the enzyme's active site observed in recent experiments.Comment: 13 pages, 10 figures (7 main text and 3 SI

Molecular dynamics studies on phenol– water clusters

Architectures of the molecules and their behavior in the clustered system are important for various consequence functions in the realistic environment. In order to gain detailed knowledge of the phenol-water clusters, 1-nano second (ns) the molecular dynamic (MD) simulation has been performed. The various structural parameters have been obtained from the MD trajectories. MD simulation reveals the presence of well-defined hydrogen-bonded network of water molecules around the phenol molecule and their dynamics. The existence of cooperativity in the hydrogen bonding and high dynamics nature of hydrogen-bonded network are evident from the present study. The calculated mutual diffusion coefficient is in close agreement with the experimental value of the phenol-water system

Precise dipole moments and quadrupole coupling constants of the cis and trans conformers of 3-aminophenol: Determination of the absolute conformation

The rotational constants and the nitrogen nuclear quadrupole coupling constants of cis-3-aminophenol and trans-3-aminophenol are determined using Fourier-transform microwave spectroscopy. We examine several $J=2\leftarrow{}1$ and $1\leftarrow{}0$ hyperfine-resolved rotational transitions for both conformers. The transitions are fit to a rigid rotor Hamiltonian including nuclear quadrupole coupling to account for the nitrogen nucleus. For cis-3-aminophenol we obtain rotational constants of A=3734.930 MHz, B=1823.2095 MHz, and C=1226.493 MHz, for trans-3-aminophenol of A=3730.1676 MHz, B=1828.25774 MHz, and C=1228.1948 MHz. The dipole moments are precisely determined using Stark effect measurements for several hyperfine transitions to $\mu_a=1.7735$ D, $\mu_b=1.5195$ D for cis-3-aminophenol and $\mu_a=0.5563$ D, $\mu_b=0.5376$ D for trans-3-aminophenol. Whereas the rotational constants and quadrupole coupling constants do not allow to determinate the absolute configuration of the two conformers, this assignment is straight-forward based on the dipole moments. High-level \emph{ab initio} calculations (B3LYP/6-31G^* to MP2/aug-cc-pVTZ) are performed providing error estimates of rotational constants and dipole moments obtained for large molecules by these theoretical methods.Comment: 9 pages, 4 tables, 3 figures (RevTeX

Molecular aggregation of thiols and alcohols: study of non-covalent interactions by microwave spectroscopy

El estudio y comprensión de las interacciones no covalentes a nivel molecular es un campo que está en continuo desarrollo y cobra vital importancia para determinar el comportamiento estructural de muchas moléculas de interés químico, tecnológico o biológico. En esta tésis doctoral se han analizado las interacciones intermoleculares implicadas en la formación de agregados moleculares neutros, tanto dímeros como productos de microsolvatación, en fase gas. Los complejos intermoleculares se han generado mediante expansiones supersónicas pulsadas, caracterizándose posteriormente mediante espectroscopía de rotación. Este trabajo ha utilizado dos técnicas espectroscópicas, incluyendo un espectrómetro de microondas con transformada de Fourier (FTMW) de tipo Balle-Flygare en el rango de frecuencias 8-20 GHz, y un espectrómetro de transformada de Fourier de banda ancha con excitación multifrecuencia (CP-FTMW) cubriendo el rango espectral de 2-8 GHz. Los complejos intermoleculares estudiados han incluido moléculas con grupos alcohol y/o tiol, con objeto de analizar las diferencias entre las interacciones intermoleculares que implican átomos de oxígeno o azufre, en especial el enlace de hidrógeno. Se han estudiado moléculas incluyendo tanto sistemas cíclicos alifáticos (ciclohexanol, ciclohexanotiol) como aromáticos (furfuril alcohol, furfuril mercaptano, tienil alcohol, tienil mercaptano). Los enlaces de hidrógeno analizados han comprendido especialmente interacciones de tipo O-H···O, O-H···S y S-H···S. La formación de los complejos intermoleculares ha revelado en algunos de ellos una gran variedad conformacional, como la observación de seis isómeros del dímero de ciclohexanol. En el caso de los monohidratos se han observado en algunos casos desdoblamientos asociados a movimientos internos de gran amplitud, como la rotación de la molécula de agua en los monohidratos de ciclohexanol y tienil mercaptano. En los casos de moléculas quirales la dimerización ha permitido observar la estabilidad relativa de los diastereoisómeros homo o heteroquirales. El estudio experimental se ha completado con diferentes cálculos teóricos de orbitales moleculares, en especial teoría del funcional de la densidad, a fin de caracterizar las interacciones estructuralmente, energéticamente y mediante análisis topológico de la densidad electrónica. El conjunto de datos experimentales y teóricos permite aumentar la información existente sobre enlaces de hidrógeno con átomos de azufre, generalmente poco estudiados, y su comparación con los análogos oxigenados.The study and understanding of non-covalent interactions at molecular level is a field in continuous development and essential to determine the structural behavior of many molecules of chemical, technological or biological interest. In this PhD thesis, the intermolecular interactions involved in the formation of neutral molecular aggregates, both dimers and microsolvation products, have been analyzed in the gas phase. The intermolecular complexes were generated by pulsed supersonic expansions, and later characterized by rotational spectroscopy. This work has used two spectroscopic techniques, including a Balle-Flygare Fourier-Transform Microwave (FTMW) spectrometer in the 8-20 GHz frequency range, and a broadband Chirped-Pulse Fourier Transform Microwave (CP-FTMW) spectrometer covering the 2-8GHz spectral range. The intermolecular complexes studied have included molecules with alcohol and / or thiol groups, in order to analyze the differences between the intermolecular interactions involving oxygen or sulfur atoms, especially hydrogen bonds. Molecules that comprise both aliphatic (cyclohexanol) and aromatic (furfuryl alcohol, furfuryl mercaptan, thenyl alcohol, thenyl mercaptan) ring systems have been studied. The analyzed hydrogen bonds included especially O-H···O, O-H···S and S-H···S interactions. The formation of intermolecular complexes has revealed a great conformational diversity in some of them, such as the observation of six isomers of the cyclohexanol dimer. With regard to the monohydrates, tunnelling splittings associated with internal large amplitude motions have been observed in some cases, such as the rotation of the water molecule in the monohydrates of cyclohexanol, thenyl alcohol and thenyl mercaptan. In the case of chiral molecules, dimerization has made it possible to observe the relative stability of homo- or heterochiral diastereoisomers. The experimental study has been supported by different theoretical molecular orbital calculations, in particular Density Functional Theory (DFT) calculations, in order to characterize the interactions structurally, energetically and by a topological analysis of electron density. The set of experimental and theoretical data will advance the existing information on hydrogen bonds involving sulfur atoms, generally scarcely studied, and their comparison with the oxygenated analogues.Departamento de Química Física y Química InorgánicaDoctorado en Físic

Hydrogen bonding in the recovery of phenols and methyl-t-butyl ether : molecular modeling and calorimetry

The purification of waste water is very important, for clean potable water is a common good and a necessity. Surface water purification is nowadays carried out on a massive industrial scale, and clean water is at our disposal virtually everywhere and always. However, cleaning industrial waste water can be a difficult task. Although apolar and slightly polar compounds can be removed from water relatively easily e.g. by extraction to an apolar phase, more polar pollutants like phenol and methyl-tert-butyl ether (MTBE), the two main compounds that this thesis deals with, cannot be removed as easily. A more effective method is therefore needed to clean water that is contaminated with either phenol or MTBE. Solvent-Impregnated Resins (SIRs) are porous polymer beads containing apolar organic extractant liquids. They are used as three-phase separation systems. When brought into contact with a SIR, a solute will preferentially partition from the aqueous phase into the impregnated solvent phase. A drawback to the use of solely the organic extraction liquid in SIRs is the limited solubility of more polar compounds like phenols and ethers in such a medium. In order to enhance extraction, complex-forming extractants can be added to the organic solvent. By means of complex formation inside the organic solvent, the overall equilibrium distribution can be shifted towards the SIR, with a concomitant enhancement of the extraction efficiency. A tight binding of the pollutant molecules to the extractant will eventually ensure high distribution coefficients. However, a moderate binding strength would enable a relatively easy regeneration of the complexing agent after a binding event, enabling multiple uses of the same compound. In this thesis hydrogen-bond (H-bond) complexation, a specific and strong yet reversible way of binding is investigated for phenol recovery and MTBE recovery from aqueous environments, involving the use of organic complexing agents that can be used inside the SIR to enhance the extraction. Potentially interesting compounds were investigated on a molecular scale by quantum chemical modeling methods and subsequent synthesis and physical characterization by primarily calorimetric means. H-bond complex formation has been evaluated and the important parameters determining the binding process have been described. After a general introduction in Chapter 1, Chapters 2 through 4 describe phenol complexation by several different classes of complexing agents. In Chapter 2, the binding of phenol and thiophenol by phosphine oxides, phosphates, and their thio-analogs was investigated. Modeling experiments, isothermal titration calorimetry (ITC) measurements, and liquid-liquid extraction experiments showed that, in principle, the binding affinity of the oxide compounds for phenol is high, whereas the sulfide compounds show only low affinity. In particular, the binding behavior of tri-n-octylphosphine oxide towards phenol and a series of electron-withdrawing group (EWG)-substituted phenols was studied, both in the presence and absence of water in the system. It was found that the presence of water in the system – as can be expected in industrial applications – yields lower binding affinities by as much as 60 %, but the binding stoichiometry remained specific and 1 : 1 complexes were still found. Electronic and steric effects were shown to play an important role in phenol binding in the investigated environment. In Chapter 3, these investigations were extended to the modeling of the full homologous series of mono-, di- and tri-substituted phosphine oxides and phosphates and their thio-analogs. Different modeling methods were used to investigate both structural and electronic elements. Dimethylphosphate was found to form the strongest complexes to the investigated phenols, but because this compound forms very strong homo-dimers in solution it cannot be used as an effective extractant. The SCS-MP2 method, that was relatively unexplored for H-bonding until now, was found to yield very accurate energy predictions, whereas the CBS-Q method was found to predict false binding affinities. Solvent effects are shown to immensely influence the binding behavior. Another, even stronger group of H-bond acceptors, amine-N-oxides, was investigated as described in Chapter 4. The binding properties for phenol and thiophenol with three different amine-N-oxides yielded very high binding affinities (up to 30 times higher than for the phosphine oxide compounds). Introduction of EWGs in the amine-N-oxides was shown to yield markedly lower binding affinities towards phenols, providing a handle to fine-tune the interaction and facilitating easier regeneration of the complexing agent in future SIR applications. Solvent effects and the influence of water in the system were investigated, and it was shown that they both influence the phenol binding strength. The results in Chapters 2 through 4 show that phosphates, phosphine oxides, and amine-N-oxides could all be used in future SIR extraction systems, and the choice between these classes of compounds can be made based on more detailed considerations. MTBE binding by several complexing agents was described in Chapter 5. A detailed modeling study of a number of different substituted phenols for MTBE binding was carried out, and the influence of solvents on the binding behavior was investigated, using a.o. the recently developed M06-2X functional and SMD solvent model. The investigated complexing agents were found to show moderate binding affinities to MTBE with binding strengths being closely linked to the acidity of the extractant. Steric effects and a proper consideration of entropic effects are also found to be important to yield successful binding of MTBE. In combination with the existing MTBE distribution coefficient for apolar phases, these moderate binding affinities were found to be able to enhance extraction, in principle, up to the point where it becomes industrially relevant to use such extractants in SIR-based extractions. Finally, in Chapter 6 the performed research is reviewed, and conclusions, recommendations and a wider perspective for future scientific challenges are given. <br/

Tautomeric Equilibria Studies by Mass Spectrometry

Tautomerism in organic chemistry has been extensively studied in condensed phase by spectrometric methods, mainly by IR and NMR techniques. Mass spectrometry studies start 40 years ago but just recently it has been recognized the importance of the mass spectral data for the study of tautomerism in the gas phase.&#xd;&#xa;Mass spectrometry can provide valuable information in regard to tautomeric equilibria when studying mass spectra among the members of different families of organic compounds.&#xd;&#xa;The relevance of the mass spectral data resides on several facts but there are two that are of key importance:&#xd;&#xa;1-&#x9;Mass spectral fragmentation assignments should be tautomer specific since the corresponding abundances ratios are supposed to be correlated to the keto/enol contents.&#xd;&#xa;2-&#x9;Ionization in the ion source is supposed to have no effect on the position of the equilibrium so that the results reflect the tautomers content in the gas phase previous to ionization.&#xd;&#xa;Some of the carbonylic compounds do not exhibit noticeable tautomerism so the fragment abundances assigned to the enol form is very low or not measurable. Since enolization is more noticeable in the case of thio-derivatives (which correlates adequately with the oxygenated analogues), the study of their mass spectra is an interesting choice to reach some degree of generalization. &#xd;&#xa;In addition, experimental findings are supported by semiempirical theoretical calculations, which probed to be adequate not only for supporting tendency correlations among the members of a compound family but also to calculate heats of tautomerization in gas phase.&#xd;&#xa;Reports using mass spectrometry for tautomerism are becoming less common. One of the reasons is that now it would appear that the interpretation of MS results is not as straightforward as it was once believed, even though in a recent review it was written that: &#x201c;Mass spectrometry is the most informative and practical method for studying and identifying tautomers in the gas phase&#x201d; [1]. &#xd;&#xa;In fact, mass spectrometry seems to be very informative for studying and identifying tautomers, because in this case external factors like solvents, intermolecular interactions, etc., can be excluded by transferring the tautomeric system into gas phase, where the process becomes truly unimolecular [1].&#xd;&#xa;This review covers the study of Tautomerism by Mass Spectrometry in the last four decades. &#xd;&#xa