Specific Water Molecules Research Articles

New challenges in quantum chemistry: Quests for accurate calculations for large molecular systems.

As quantum chemistry plays a more and more central role in many complicated chemical problems, it has become necessary to obtain accurate results for large molecular systems. Conventional quantum chemistry methods are either too expensive to apply to large systems or too approximate for the results to be reliable, and they fail to satisfy this requirement. A variety of different approaches is being developed with the aim of achieving this goal: local correlation methods; divide-and-conquer methods; linear-scaling density functional methods based on the fast multipole and other approximations; effective potential methods; and hybrid methods. ONIOM (our N-layered integrated molecular orbital plus molecular mechanics method), developed by the authors, is a hybrid method in which a large molecular system is divided into onion-skin-like layers, and different quantum chemistry/molecular mechanics methods are used for different parts of the system; the results are combined to extrapolatively estimate the results of high-level calculation for the real system. Several applications of ONIOM will be discussed.

Interaction of internal water molecules with the schiff base in the L intermediate of the bacteriorhodopsin photocycle.

In the photocycle of bacteriorhodopsin (BR), the first proton movement, from the Schiff base to Asp85, occurs after the formation of the L intermediate. In L, the C [double bond] N bond of the Schiff base is strained, and the nitrogen interacts strongly with its counterion. The present study seeks to detect the interaction of internal water molecules with the Schiff base in L using difference FTIR spectroscopy at 170 K. The coupled modes of the hydrogen-out-of plane bending vibrations (HOOPs) of the N-H and C(15)-H of the protonated Schiff base are detected as a broad band centered at 911 cm(-1) for BR. A set of bands at 1073, 1064, and 1056 cm(-1) for L is shown to arise from the coupling of the HOOP with the overtones of interacting water O-H vibrations. Interaction with water was shown by the decreased intensity of the HOOPs of L in H(2)(18)O and by the influence of mutants that have been shown to perturb specific internal water molecules in BR. In contrast, the HOOP band of initial BR was not affected by these mutations. In D85N, the coupled HOOP of BR is depleted, while the coupled HOOPs of L are shifted. The results indicate that the Schiff base interacts with water in the L state but in a different manner than in the BR state. Moreover, the effects of mutations suggest that cytoplasmic water close to Thr46 (Wat46) either interacts stronger with the Schiff base in L or that it is important in stabilizing another water that does.

Sensitivity of molecular docking to induced fit effects in influenza virus neuraminidase.

Many proteins undergo small side chain or even backbone movements on binding of different ligands into the same protein structure. This is known as induced fit and is potentially problematic for virtual screening of databases against protein targets. In this report we investigate the limits of the rigid protein approximation used by the docking program, GOLD, through cross-docking using protein structures of influenza neuraminidase. Neuraminidase is known to exhibit small but significant induced fit effects on ligand binding. Some neuraminidase crystal structures caused concern due to the bound ligand conformation and GOLD performed poorly on these complexes. A 'clean' set, which contained unique, unambiguous complexes, was defined. For this set, the lowest energy structure was correctly docked (i.e. RMSD < 1.5 A away from the crystal reference structure) in 84% of proteins, and the most promiscuous protein (1mwe) was able to dock all 15 ligands accurately including those that normally required an induced fit movement. This is considerably better than the 70% success rate seen with GOLD against general validation sets. Inclusion of specific water molecules involved in water-mediated hydrogen bonds did not significantly improve the docking performance for ligands that formed water-mediated contacts but it did prevent docking of ligands that displaced these waters. Our data supports the use of a single protein structure for virtual screening with GOLD in some applications involving induced fit effects, although care must be taken to identify the protein structure that performs best against a wide variety of ligands. The performance of GOLD was significantly better than the GOLD implementation of ChemScore and the reasons for this are discussed. Overall, GOLD has shown itself to be an extremely good, robust docking program for this system.

Protein-protein recognition: an experimental and computational study of the R89K mutation in Raf and its effect on Ras binding.

Binding of the protein Raf to the active form of Ras promotes activation of the MAP kinase signaling pathway, triggering cell growth and differentiation. Raf/Arg89 in the center of the binding interface plays an important role determining Ras-Raf binding affinity. We have investigated experimentally and computationally the Raf-R89K mutation, which abolishes signaling in vivo. The binding to [gamma-35S]GTP-Ras of a fusion protein between the Raf-binding domain (RBD) of Raf and GST was reduced at least 175-fold by the mutation, corresponding to a standard binding free energy decrease of at least 3.0 kcal/mol. To compute this free energy and obtain insights into the microscopic interactions favoring binding, we performed alchemical simulations of the RBD, both complexed to Ras and free in solution, in which residue 89 is gradually mutated from Arg into Lys. The simulations give a standard binding free energy decrease of 2.9+/-1.9 kcal/mol, in agreement with experiment. The use of numerous runs with three different force fields allows insights into the sources of uncertainty in the free energy and its components. The binding decreases partly because of a 7 kcal/mol higher cost to desolvate Lys upon binding, compared to Arg, due to better solvent interactions with the more concentrated Lys charge in the unbound state. This effect is expected to be general, contributing to the lower propensity of Lys to participate in protein-protein interfaces. Large contributions to the free energy change also arise from electrostatic interactions with groups up to 8 A away, namely residues 37-41 in the conserved effector domain of Ras (including 4 kcal/mol from Ser39 which loses a bifurcated hydrogen bond to Arg89), the conserved Lys84 and Lys87 of Raf, and 2-3 specific water molecules. This analysis will provide insights into the large experimental database of Ras-Raf mutations.

Binding of a substrate analog to a domain swapping protein: X-ray structure of the complex of bovine seminal ribonuclease with uridylyl(2',5')adenosine.

Bovine seminal ribonuclease (BS-RNase) is a unique member of the pancreatic-like ribonuclease superfamily. The native enzyme is a mixture of two dimeric forms with distinct structural features. The most abundant form is characterized by the swapping of N-terminal fragments. In this paper, the crystal structure of the complex between the swapping dimer and uridylyl(2',5')adenosine is reported at 2.06 A resolution. The refined model has a crystallographic R-factor of 0.184 and good stereochemistry. The quality of the electron density maps enables the structure of both the inhibitor and active site residues to be unambiguously determined. The overall architecture of the active site is similar to that of RNase A. The dinucleotide adopts an extended conformation with the pyrimidine and purine base interacting with Thr45 and Asn71, respectively. Several residues (Gln11, His12, Lys41, His119, and Phe120) bind the oxygens of the phosphate group. The structural similarity of the active sites of BS-RNase and RNase A includes some specific water molecules believed to be relevant to catalytic activity. Upon binding of the dinucleotide, small but significant modifications of the tertiary and quaternary structure of the protein are observed. The ensuing correlation of these modifications with the catalytic activity of the enzyme is discussed.

Thermal Denaturation of Ribonuclease A Characterized by Water 17O and 2H Magnetic Relaxation Dispersion

Water oxygen-17 and deuteron nuclear magnetic relaxation dispersion (NMRD) measurements were used to characterize ribonuclease A (RNase A) in the course of thermal denaturation at pH 2 and 4. The structure and dynamics of the protein were probed by specific long-lived water molecules, by the short-lived surface hydration, and by labile side-chain hydrogens. The NMRD data show that native RNase A contains at least three water molecules with a mean residence time of 8 ns at 27 degreesC and an activation enthalpy of ca. 40 kJ mol-1. These water molecules are identified with some or all of six ordered water molecules partly buried in surface pockets in the crystal structure of RNase A. The loss of the 17O dispersion at higher temperatures demonstrates that, in the thermally denatured protein, these surface pockets are either not present or undergoing large structural fluctuations on a subnanosecond time scale. The relaxation dispersion step vanishes monotonically and essentially in concert with the CD denaturation curves, thus ruling out the existence of equilibrium intermediates with a substantial amount of non-native and long-lived hydration water. The NMRD data show that thermally denatured RNase A has a relatively compact but highly flexible structure. The global solvent exposure and the hydrodynamic volume of the denatured protein are much less than for maximally unfolded disulfide-intact RNase A. The NMRD data show that thermal denaturation is accompanied by a large reduction of the mean-square orientational order parameter of side-chain O-H bonds, implying that, in the denatured state, these side chains sample a wide distribution of conformational states on a subnanosecond time scale.

Molecular Modeling of the Enantioselectivity in Lipase-Catalyzed Transesterification Reactions

Two strategies based on the use of subsets for calculating the enantioselectivity in lipase-catalyzed transesterifications using the CHARMM force field were investigated. Molecular dynamics was used in our search for low energy conformations. Molecular mechanics was used for refining these low energy conformations. A tetrahedral intermediate with a rigid central part was used for mimicking the transition state. The energy differences between the transition states of the diastereomeric enzyme-substrate complexes were calculated. The way of defining the subsets was based on two fundamentally different strategies. The first strategy used predefined parts of the enzyme and the substrate as subsets. The second approach formed energy-based subsets, varying in size with the substrates studied. The selection of residues to be included in these energy-based subsets was based on the energy of the interaction between the specific residue or water molecule and the transition state. The reaction studied was the kinetic resolution of secondary alcohols in transesterifications using the Candida antarctica lipase B as chiral biocatalyst. The secondary alcohols used in the study were 2-butanol, 3-methyl-2-butanol, and 3,3-dimethyl-2-butanol.

Interfacial Behavior of Ionophoric Systems: Molecular Dynamics Studies on 18-Crown-6 and Its Complexes at the Water-Chloroform Interface

We report a series of molecular dynamics simulations on 18-crown-6 (18C6) uncomplexed, and on its K+Pic- complexes, at the water-chloroform interface, using an explicit representation of the solvents. The (18C6)10 aggregate, initially at the interface, displays after 1 ns a complex equilibrium between molecules adsorbed at the interface, and others which have diffused to the organic phase. The (18C6·K+Pic-)6 inclusive complexes, initially at the interface, also display after 1 ns, an equilibrium between 18C6 molecules, 18C6·K+ complexes and the Pic- counterions adsorbed at the interface, while some decomplexed K+ cations are captured by water. The spontaneous migration of a 18C6·K+ complex to the organic phase is also observed, facilitated by specific water molecules and the Pic- counterions. Comparison with simulations on the (18C6·K+Pic-)1 and (18C6·Sr2+2Pic-)1 complexes highlights the effect of concentration and of the cationic charge on the interfacial behavior. We finally describe a “computer extraction experiment”, which starts with 18C6 molecules in the organic phase and K+Pic- ion pairs in water. The results are discussed in relation with experimental data. We emphasize the surfactant-like behavior of free and complexed extractant molecules, the preorganization of ionophores induced by the interface, and the role of counterions. Surface active anions attract the cations at the interface and facilitate their capture by the adsorbed ionophores. Synergistic effects should result from the adsorption of co-extractant molecules or/and of counterions at the interface.

Analysis of shorthorn sculpin antifreeze protein stereospecific binding to (2–1 0) faces of ice

In this paper we report the results of our studies on the stereospecific binding of shorthorn sculpin antifreeze protein (AFP) to (2 -1 0) secondary prism faces of ice. Using ice crystal growth and etching techniques together with molecular modeling, molecular dynamics, and energy minimization, we explain the nature of preferential binding of shorthorn sculpin AFP along the [1 2 2] direction on (2- 1 0) planes. In agreement with ice etching studies, the mechanism of preferential binding suggested by molecular modeling explains why the binding of shorthorn sculpin AFP occurs along [1 2 2] and not along its mirror symmetry-related direction [-1 -2 2] on (2 -1 0). This binding mechanism is based on the protein-crystal surface enantioselective recognition that utilizes both alpha-helical protein backbone matching to the (2 -1 0) surface topography and matching of side chains of polar/charged residues with specific water molecule positions in the ice surface. The mechanisms of winter flounder and shorthorn sculpin antifreeze binding to ice are compared.

AM1-SM2 and PM3-SM3 parameterized SCF solvation models for free energies in aqueous solution.

Two new continuum solvation models have been presented recently, and in this paper they are explained and reviewed in detail with further examples. Solvation Model 2 (AM1-SM2) is based on the Austin Model 1 and Solvation Model 3 (PM3-SM3) on the Parameterized Model 3 semiempirical Hamiltonian. In addition to the incorporation of phosphorus parameters, both of these new models address specific deficiencies in the original Solvation Model 1 (AM1-SM1), viz., (1) more accurate account is taken of the hydrophobic effect of hydrocarbons, (2) assignment of heavy-atom surface tensions is based on the presence or absence of bonded hydrogen atoms, and (3) the treatment of specific hydration-shell water molecules is more consistent. The new models offer considerably improved performance compared to AM1-SM1 for neutral molecules and essentially equivalent performance for ions. The solute charges within the Parameterized Model 3 Hamiltonian limit the utility of PM3-SM3 for compounds containing nitrogen and possibly phosphorus. For other systems both AM1-SM2 and PM3-SM3 give realistic results, but AM1-SM2 in general outperforms PM3-SM3. Key features of the models are discussed with respect to alternative approaches.

Liquid Phase Kinetic and Thermodynamic Studies on Promoted Nickel Catalysts used for H/D Exchange

Successful choice of the suitable catalyst composition for hydrogen/deuterium exchange reaction between hydrogen and water requires understanding of chemical kinetics as well as surface properties and activity in both liquid- and vapour phases. The present study deals with the thermodynamic and kinetic behaviour of a simple liquid phase reaction on the surface of nickel catalyst promoted with zirconium oxide used for the H/D exchange. This study helps understand the condensation of water on the catalyst surface during operation which leads to its poisoning. In this specific case water molecules are responsible for selective poisoning of this type of catalysts. The reaction rate constant, the increase in Gibb,s free energy, the enthalpy, the entropy of activation and the energy of activation were calculated.

Possible role for water dissociation in the slow binding of phosphorus-containing transition-state-analogue inhibitors of thermolysin.

A number of phosphonamidate and phosphonate tripeptide analogues have been studied as transition-state-analogue inhibitors of the zinc endopeptidase thermolysin. Those with the form Cbz-GlyP(Y)Leu-X [ZGP(Y)LX, X = NH2 or amino acid, Y = NH or O linkage] are potent (Ki = 9-760 nM for X = NH, 9-660 microM for X = O) but otherwise ordinary in their binding behavior, with second-order rate constants for association (kon) greater than 10(5) M-1 s-1. Those with the form Cbz-XP(Y)-Leu-Ala [ZXP(Y)LA,XP = alpha-substituted phosphorus amino acid analogue] are similarly potent (Ki for ZFPLA = 68 pM) but slow binding (kon less than or equal to 1300 M-1 s-1). Several kinetic mechanisms for slow binding behavior are considered, including two-step processes and those that require prior isomerization of inhibitor or enzyme to a rare form. The association rates of ZFPLA and ZFP(O)LA are first order in inhibitor concentration up to 1-2 mM, indicating that any loose complex along the binding pathway must have a dissociation constant above this value. The crystallographic investigation described in the preceding paper [Holden, H. M., Tronrud, D. E., Monzingo, A. F., Weaver, L. H., & Matthews, B. W. (1987) Biochemistry (preceding paper in this issue)] identifies a specific water molecule in the active site that may hinder binding of the alpha-substituted inhibitors. The implication of this observation for a mechanism for slow binding is discussed.

An Analysis of the Enzyme-Inhibitor Binding Interactions for Phosphonic Acid Transition State Analogs of Thermolys in

Abstract Potent inhibitors of the zinc endopeptidase thermolysin are produced on replacement of the scissile peptide linkage with phosphonamidate or phosphonate ester moieties. These inhibitors have been shown to be transition state analogs, and a comparison between the esters and amidates reveals the intrinsic binding energy due to a specific hydrogen bonding interaction. Incorporation of an α-substituted phosphonic acid analog leads to slow-binding behavior on the part of the inhibitors, which is attributed to expulsion of a specific water molecule from the active site.

Pulse radiolysis study on electrons trapped in semiclathrates and non-clathrate hydrates

Trapping of electrons in specific water molecule vacancies, observed previously in crystalline aqueous clathrates, has also been found in semiclathrates (e.g., tetramethylammonium hydroxide pentahydrate), in clathrates showing hydrogen bonds between host and guest molecules (e.g., piperazine clathrate), and also in inorganic hydrates (e.g., sodium carbonate decahydrate). The lifetime of the electron is sometimes longer than in the case of true clathrates; e.g., t/sub 1/2/ = 3.5 ms (first-order decay in piperazine clathrate). The existence of comparatively long-lived electrons at room temperature may be considered a general phenomenon. The condition for its occurrence is the presence of OH/sup -/ or F/sup -/ anion, which can substitute for H/sub 2/O in the aqueous part of the compound and when displaced leaves an electron trap. In other ionic and nonionic compounds, the condition for the trapping of long-lived electrons is protonation of the principal compound, thereby leaving the solution or melt alkaline during the crystallization of the hydrate. Interpretation in terms of preexistent traps invokes the crystal imperfections chemistry, which in the case of hydrates has not yet been noticed. It has been assumed that electrons occupy the vacancies temporarily revealing their presence. In some clathrates and other hydrates, the long-lived electron trapsmore » do not occur; instead, the electron shows a similar spectrum (620 nm maximum), decaying by 2-3 orders of magnitude faster than in long-lived traps. It is assumed that this is a case of electron digging its own hole, although an alternative explanation may be through trapping in other kinds of crystal imperfections in the aqueous moiety of the hydrate.« less

Muscle contraction and enzymatic action: Challenges to quantum chemistry

The energy of muscle contraction is generated by interactions between the proteins actin and myosin, acting together to hydrolyze a metaphosphate bond of ATP in the presence of Mg++, using the oxygen of a specific water molecule. The molecular setting for these events is described. A key feature is identified to be an attack by the water oxygen on one of the POP bonds. The resulting hydrolysis liberates a phosphate group which is forcefully repelled from its original setting, perturbing metal ions and electron orbitals in the vicinity. The hypothesis is advanced that the accompanying perturbations excite an electron cloud to a hgher energy level, leading to a forceful expansion of the electron cloud. A model is presented representing the S-l head of myosin as being tilted with reference to the coordinates of the actin filament by the expansion of this cloud. This tilt represents a leverlike movement of the S-1 myosin head which is capable of powering the work of muscle contraction. Since the most important molecular participants in muscle contraction are present in all cells, the issues raised here are of very general biological significance.

Nuclear spin decoupling in ENDOR spectroscopy

The decoupling of dipolar proton-proton interactions by a strong second radio frequency field in ENDOR spectroscopy is reported. A theoretical description of the decoupling effect for a system with one electron and two nuclear spins of ½ is given for anisotropic g and hyperfine tensors. The results indicate that the hyperfine tensor of the pumped nucleus influences the decoupling behaviour remarkably. Transition frequencies and intensities predicted by theory are compared with experimental data from Cu(gly)2 in α-glycine. Further applications of this spin decoupling technique are discussed. As an example, the assignment of ENDOR proton lines to specific water molecules in Cu2+ : (NH4)2Mg(SO4)2·6H2O is presented.