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The dynamics of processes relevant to chemistry and biophysics on rough free energy landscapes is investigated using a recently developed algorithm to solve the Smoluchowski equation. Two different processes are considered: ligand rebinding in MbCO and protein folding. For the rebinding dynamics of carbon monoxide (CO) to native myoglobin (Mb) from locations around the active site, the two-dimensional free energy surface (FES) is constructed using extensive molecular dynamics simulations. The surface describes the minima in the A state (bound MbCO), CO in the distal pocket and in the Xe4 pocket, and the transitions between these states and allows to study the diffusion of CO in detail. For the folding dynamics of protein G, a previously determined two-dimensional FES was available. To follow the diffusive dynamics on these rough free energy surfaces, the Smoluchowski equation is solved using the recently developed hierarchical discrete approximation method. From the relaxation of the initial nonequilibrium distribution, experimentally accessible quantities such as the rebinding time for CO or the folding time for protein G can be calculated. It is found that the free energy barrier for CO in the Xe4 pocket and in the distal pocket (B state) closer to the heme iron is approximately 6 kcal/mol which is considerably larger than the inner barrier which separates the bound state and the B state. For the folding of protein G, a barrier of approximately 10 kcal/mol between the unfolded and the folded state is consistent with folding times of the order of milliseconds.

Fully atomistic simulations are used to characterize the molecular dynamics (MD) of acridine orange (3,6-dimethylaminoacridine) at a chromatographic interface. Multiple 1 ns MD simulations were performed for acridine orange at the interface between three different acetonitrile/water mixtures (0/100, 20/80, and 50/50) with C8 and C18 alkyl chains. The diffusion coefficient, D, of acridine orange in pure solvent was found to be 4 times smaller at the water/C18 interface (D = 0.022 x 10(-4) cm2/s) than in bulk water (D = 0.087 x 10(-4) cm2/s), in qualitative agreement with experiment. Rotational reorientation times were 20 and 700 ps, which also agree favorably with the measured time scales of 130 and 740 ps. Contrary to experiment, the simulations found that for increasing surface coverage, the diffusion coefficient for acridine decreased. Detailed analysis of the solvent structure showed that the transport properties of acridine were primarily governed by the solvent distribution above the functionalized surface. The solvent structure, in turn, was largely determined by the surface consisting of the silica layer, the alkyl chains, and their functionalization.

The vibrational dynamics of the cyanide anion and the heme group in MbCN (CN complexed to Myoglobin) are investigated using molecular dynamics simulations. A previously calculated quantum-chemical heme-ligand potential-energy surface together with a three-center charge model for the iron-ligand center that captures both polarization and ligand-to-metal charge transfer allows for a detailed description of the interactions around the active site. It is found that the CN binding orientation (Fe--CN or Fe--NC) to the heme affects the stretching frequency of the ligand, with a 25 cm-1 difference in the fundamental wavenumber between the two orientations as well as a change in bond length. The charge model also captures such crucial interactions as the possible hydrogen bond between the ligand and the His64 residue. This interaction is weakened when the ligand binds in the Fe--NC conformation but is also sensitive to the protonation state of His64. The structural changes around the active site, the observation of water penetration for the Fe--NC conformation, the computed IR spectrum, and the energetics suggest that the Fe--CN conformation with Hisepsilon64 is the most likely one. The water accessibility of the active site is also found to be related to the protonation state of His64. The presence of water in the active site could also affect the IR band of the C--N stretch mode. Thus, IR spectroscopy of the C--N stretch is a potentially useful reporter of ligand isomers and active-site structure.

The binding of NO to iron is involved in the biological function of many heme proteins. Contrary to ligands like CO and O(2), which only bind to ferrous (Fe(II)) iron, NO binds to both ferrous and ferric (Fe(III)) iron. In a particular protein, the natural oxidation state can therefore be expected to be tailored to the required function. Herein, we present an ab initio potential-energy surface for ferric iron interacting with NO. This potential-energy surface exhibits three minima corresponding to eta(1)-NO coordination (the global minimum), eta(1)-ON coordination and eta(2) coordination. This contrasts with the potential-energy surface for Fe(II)-NO, which exhibits only two minima (the eta(2) coordination mode for Fe(II) is a transition state, not a minimum). In addition, the binding energies of NO are substantially larger for Fe(III) than for Fe(II). We have performed molecular dynamics simulations for NO bound to ferric myoglobin (Mb(III)) and compare these with results obtained for Mb(II). Over the duration of our simulations (1.5 ns), all three binding modes are found to be stable at 200 K and transiently stable at 300 K, with eventual transformation to the eta(1)-NO global-minimum conformation. We discuss the implication of these results related to studies of rebinding processes in myoglobin.

The energetics, dynamics, and infrared spectroscopy of the shared proton in different chemical environments is investigated using molecular dynamics simulations. A three-dimensional potential energy surface (PES) suitable for describing proton transfer between an acceptor and a donor oxygen atom is combined with an all-atom force field to carry out reactive molecular dynamics simulations. The construction of the fully dimensional PES is inspired from the established mixed quantum mechanics/molecular mechanics treatment of larger systems. The "morphing potential" method is used to transform the generic PES for proton transfer along an O...H+...O motif into a three-dimensional PES for proton transfer in protonated diglyme. Using molecular dynamics simulations at finite temperature, the gas phase infrared spectra are calculated for both species from the Fourier transform of the dipole moment autocorrelation function. For protonated diglyme the modes involving the H+ motion are strongly mixed with other degrees of freedom. At low temperature, the O...H+...O asymmetric stretching vibration is found at 870 cm-1, whereas for H5O2+ this band is at 724 cm-1. As expected, the vibrational bands of protonated diglyme show no temperature dependence whereas for H5O2+ at T = 100 K the proton transfer mode is found at 830 cm-1, in good agreement with 861 cm-1 from very recent molecular dynamics simulations.

The dynamics of the cyanide anion bound to sperm-whale myoglobin is investigated using atomistic simulations. With density-functional theory, a 2D potential energy surface for the cyanide-heme complex is calculated. Two deep minima with a stabilization energy of approximately 50 kcal/mol corresponding to two different binding orientations (Fe-CN and Fe-NC) of the ligand are found. The Fe-CN conformation is favored over Fe-NC by several kcal/mol. Mixed quantum mechanics/molecular mechanics calculations show that the binding orientation affects the bond strength of the ligand, with a significantly different bond length and a 25 cm-1 shift in the fundamental CN-frequency. For the molecular dynamics (MD) simulations, a 3-center fluctuating charge model for the Fe-CN unit is developed that captures polarization and ligand-metal charge transfer. Stability arguments based on the energetics around the active site and the CN- frequency shifts suggest that the Fe-CN conformation with epsilon-protonation of His epsilon 64 are most likely, which is in agreement with experiment. Both equilibrium and nonequilibrium MD simulations are carried out to investigate the relaxation time scale and possible relaxation pathways in bound MbCN. The nonequilibrium MD simulations with a vibrationally excited ligand reveal that vibrational relaxation takes place on a time scale of hundreds of picoseconds within the active site. This finding supports the hypothesis that the experimentally observed relaxation rate (3.6 ps) reflects the repopulation of the electronic ground state.

As previously shown, alkali and alkaline earth metal iodides in nonaqueous, aprotic solvents behave like transition metal halides, forming cis- and trans-dihalides with various neutral O-donor ligands. These compounds can be used as precursors for the synthesis of new mixed alkali/alkaline earth metal aggregates. We show here that Ln2+ ions form isostructural cluster compounds. Thus, with LiOtBu, 50% of the initial iodide can be replaced in MI2, M=Ca, Sr, Ba, Eu, to generate the mixed-metal alkoxide aggregates [IM(OtBu)4{Li(thf)}4(OH)], for which the M--OH contacts were investigated by theoretical methods. With M'OPh (M'=Li, Na), a new mixed-metal aryloxide cluster type [MM'6(OPh)8(thf)6] is obtained for M=Ca, Sr, Ba, Sm, Eu. Their stability versus DME (DME=1,2-dimethoxyethane) as bidentate ligand is studied.

A new computational approach is proposed to probe the importance of residue side chains for the stability of a protein fold. Computational mutations to estimate protein stability (CMEPS) is based on the notion that the binding free energy corresponding to the complexation of a given side chain, considered as a "pseudo-ligand" of the wild type protein, reflects the importance of this side chain to the thermodynamic stability of the protein. The contribution of a particular side chain to the folding energy is estimated according to the molecular mechanics-generalized born surface area MM-GBSA approach, using a single molecular dynamics simulation trajectory of the wild type protein. CMEPS is a first principles method which does not contain any adjustable parameter that could be fitted to experimental data. The approach is first validated for Barnase and the B1 domain of protein L, for which a correlation coefficient R = 0.73, between experimental and CMEPS calculated DeltaDeltaG values, is found and then applied to the insulin monomer. In the present application, CMEPS replaces each amino acid by an alanine residue. Therefore, most mutations lead to cavities in the protein. From this the change in stability can be correlated with increased cavity volume. For insulin, this correlation is very similar compared with data previously analyzed for T4 lysozyme from an experiment for buried apolar side chains. There, the increased cavity volume has been related to the hydrophobic effect. However, since CMEPS uses the energetics in terms of electrostatic and van der Waals interactions (and not the hydrophobic effect which is difficult to relate to physical interactions), it is possible to study the effect of mutations of polar and solvent accessible side chains. According to CMEPS, residues Leu A16, Tyr A19, Leu B11, Leu B15, and Arg B22 are most important for the stability of the monomeric insulin fold. This is in agreement with experimental data. As a consequence, mutation of these residues may lead to misfolded and inactive insulin analogues., (Copyright 2006 Wiley Periodicals, Inc.)

The influence of possible inaccuracies that can arise during homology modeling of protein structures used for ligand binding studies were investigated with the molecular mechanics generalized Born surface area (MM-GBSA) method. For this, a family of well-characterized HIV-I protease-inhibitor complexes was used. Validation of MM-GBSA led to a correlation coefficient ranging from 0.72 to 0.93 between calculated and experimental binding free energies DeltaG. All calculated DeltaG values were based on molecular dynamics simulations with explicit solvent. Errors introduced into the protein structure through misplacement of side-chains during rotamer modeling led to a correlation coefficient between DeltaG(calc) and DeltaG(exp) of 0.75 compared with 0.90 for the correctly placed side chains. This is in contrast to homology models for members of the retroviral protease family with template structures ranging in sequence identity between 32% and 51%. For these protein models, the correlation coefficients vary between 0.84 and 0.87, which is considerably closer to the original protein (0.90). It is concluded that HIV-I low sequence identity with the template structure still allows creating sufficiently reliable homology models to be used for ligand-binding studies, although placement of the rotamers is a critical step during the modeling., ((c) 2006 Wiley-Liss, Inc. V)

Cyclic di-guanosine monophosphate is a bacterial second messenger that has been implicated in biofilm formation, antibiotic resistance, and persistence of pathogenic bacteria in their animal host. Although the enzymes responsible for the regulation of cellular levels of c-di-GMP, diguanylate cyclases (DGC) and phosphodiesterases, have been identified recently, little information is available on the molecular mechanisms involved in controlling the activity of these key enzymes or on the specific interactions of c-di-GMP with effector proteins. By using a combination of genetic, biochemical, and modeling techniques we demonstrate that an allosteric binding site for c-di-GMP (I-site) is responsible for non-competitive product inhibition of DGCs. The I-site was mapped in both multi- and single domain DGC proteins and is fully contained within the GGDEF domain itself. In vivo selection experiments and kinetic analysis of the evolved I-site mutants led to the definition of an RXXD motif as the core c-di-GMP binding site. Based on these results and based on the observation that the I-site is conserved in a majority of known and potential DGC proteins, we propose that product inhibition of DGCs is of fundamental importance for c-di-GMP signaling and cellular homeostasis. The definition of the I-site binding pocket provides an entry point into unraveling the molecular mechanisms of ligand-protein interactions involved in c-di-GMP signaling and makes DGCs a valuable target for drug design to develop new strategies against biofilm-related diseases.

One of the predominant aims of insulin therapy for diabetes is to appropriately mimic physiological insulin secretion levels and their correlation with glucose concentration in healthy individuals. This report outlines current methods and their limitations in glycemic control and their possible relationship to insufficient knowledge about the structure and dynamics of the insulin hormone itself. Based on recent experimental and computational work, a possible approach to less-invasive insulin administration is sketched.

An experimental and theoretical study of the pressure broadening and the pressure shift of three HCO(+) rotational lines (j=4<--3, 5<--4 and 6<--5) perturbed by collisions with Ar is presented. The measurements are carried out at 77 K and are compared to close-coupling calculations performed on an accurate potential energy surface for the Ar-HCO(+) interaction extending from small to very large separations between the ion and the perturber. For the pressure broadening, agreement between experiment and theory is satisfactory for both close-coupling and semiclassical calculations. For the pressure shift, however, close-coupling calculations are superior. The results agree with experiment in sign and order of magnitude, while semiclassical calculations are inaccurate for the shift of the presently studied lines because they neglect the contribution of strong collisions.

A new surface-crossing algorithm suitable for describing bond-breaking and bond-forming processes in molecular dynamics simulations is presented. The method is formulated for two intersecting potential energy manifolds which dissociate to different adiabatic states. During simulations, crossings are detected by monitoring an energy criterion. If fulfilled, the two manifolds are mixed over a finite number of time steps, after which the system is propagated on the second adiabat and the crossing is carried out with probability one. The algorithm is extensively tested (almost 0.5 mus of total simulation time) for the rebinding of NO to myoglobin. The unbound surface (Fe...NO) is represented using a standard force field, whereas the bound surface (Fe-NO) is described by an ab initio potential energy surface. The rebinding is found to be nonexponential in time, in agreement with experimental studies, and can be described using two time constants. Depending on the asymptotic energy separation between the manifolds, the short rebinding timescale is between 1 and 9 ps, whereas the longer timescale is about an order of magnitude larger. NO molecules which do not rebind within 1 ns are typically found in the Xenon-4 pocket, indicating the high affinity of NO to this region in the protein.

The structures and infrared spectra of protonated ammonia clusters NH(4+)(NH3)n, for n < or = 8, are investigated using density functional-theory (DFT) calculations and semiempirical DFT/molecular dynamics simulations. For n < 5 the clusters are found to be mostly stable up to 100 K, while the larger clusters (n > or = 5) isomerize. Temperature effects are taken into account by performing ab initio molecular dynamics simulations with the computationally tractable self-consistent charges density functional tight-binding method. The infrared spectra at 10 K for the most stable isomers for n = 3-8 compare qualitatively with predissociation experiments, and using a common scaling factor almost quantitative agreement is found. For n > or = 6 the notion of multiple isomers present under the experimental conditions is supported. Of the 13 stable structures for n = 8 only three are found to survive at 100 K. All other clusters isomerize. Cluster structures are inferred from the analysis of the cumulative radial distribution function of the ammonia molecules surrounding the NH(4+) core. The infrared spectra are found to be typical for the structure of the clusters, which should help to relate the experimentally measured infrared spectra to the number and identity of the contributing isomers. For clusters that reorganize to a more stable isomer during the dynamics, the infrared spectrum is generally similar to that of the stable isomer itself. The clusters are found to preferably form globular structures, although chain-like arrangements are also among the low-energy configurations.

Ligands such as CO, O(2), or NO are involved in the biological function of myoglobin. Here we investigate the energetics and dynamics of NO interacting with the Fe(II) heme group in native myoglobin using ab initio and molecular dynamics simulations. At the global minimum of the ab initio potential energy surface (PES), the binding energy of 23.4 kcal/mol and the Fe-NO structure compare well with the experimental results. Interestingly, the PES is found to exhibit two minima: There exists a metastable, linear Fe-O-N minimum in addition to the known, bent Fe-N-O global minimum conformation. Moreover, the T-shaped configuration is found to be a saddle point, in contrast to the corresponding minimum for NO interacting with Fe(III). To use the ab initio results for finite temperature molecular dynamics simulations, an analytical function was fitted to represent the Fe-NO interaction. The simulations show that the secondary minimum is dynamically stable up to 250 K and has a lifetime of several hundred picoseconds at 300 K. The difference in the topology of the heme-NO PES from that assumed previously (one deep, single Fe-NO minimum) suggests that it is important to use the full PES for a quantitative understanding of this system. Why the metastable state has not been observed in the many spectroscopic studies of myoglobin interacting with NO is discussed, and possible approaches to finding it are outlined.

A calculation of the binding free energy for the dimerization of insulin has been performed using the molecular mechanics-generalized Born surface area approach. The calculated absolute binding free energy is -11.9 kcal/mol, in approximate agreement with the experimental value of -7.2 kcal/mol. The results show that the dimerization is mainly due to nonpolar interactions. The role of the hydrogen bonds between the 2 monomers appears to give the direction of the interactions. A per-atom decomposition of the binding free energy has been performed to identify the residues contributing most to the self association free energy. Residues B24-B26 are found to make the largest favorable contributions to the dimerization. Other residues situated at the interface between the 2 monomers were found to make favorable but smaller contributions to the dimerization: Tyr B16, Val B12, and Pro B28, and to an even lesser extent, Gly B23. The energy decomposition on a per-residue basis is in agreement with experimental alanine scanning data. The results obtained from a single trajectory (i.e., the dimer trajectory is also used for the monomer analysis) and 2 trajectories (i.e., separate trajectories are used for the monomer and dimer) are similar., ((c) 2005 Wiley-Liss, Inc.)

The rebinding of CO to myoglobin (Mb) from locations around the active site is studied using a combination of molecular dynamics and stochastic simulations for native and L29F mutant Mb. The interaction between the dissociated ligand and the protein environment is described by the recently developed fluctuating three-point charge model for the CO molecule. Umbrella sampling along trajectories, previously found to sample the binding site (B) and the Xe4 pocket, is used to construct free-energy profiles for the ligand escape. On the basis of the Smoluchowski equation, the relaxation of different initial population distributions is followed in space and time. For native Mb at room temperature, the calculated rebinding times are in good agreement with experimental values and give an inner barrier of 4.3 kcal/mol between the docking site B (Mb...CO) and the A state (bound MbCO), compared to an effective barrier, Heff, of 4.5 kcal/mol and barriers into the majority conformation A1 and the minority conformation A3 of 2.4 and 4.3 kcal/mol, respectively. In the case of the L29F mutant, the free-energy surface is flatter and the dynamics is much more rapid. As was found in experiment, escape to the Xe4 pocket is facile for L29F whereas, for native Mb, the barriers to this site are larger. At lower temperatures, the rebinding dynamics is delayed by orders of magnitude also due to increased barriers between the docking sites.

We present an efficient and numerically robust algorithm to follow diffusive processes on rough potential energy surfaces. The hierarchical nature of the algorithm (hierarchical discrete approximation or HDA) fully explores the fine- and coarse-grained structure of the underlying interaction potential. The present approach does not impose any restriction on the topology of the potential. The hierarchical grid allows to capture the roughness of the potential and achieve significant reduction of computational time using fewer grid points compared to other DA methods. HDA is shown to be accurate and efficient by comparing with results from the conventional DA and from the "mean first passage time" (MFPT) method. Using potential-optimized grids HDA monotonically converges to results from an analytical treatment for a very rough interaction potential (107 minima). Contrary to MFPT the solution from HDA is numerically stable. Because of the hierarchical structure of the method HDA can be extended to multidimensional problems.

We present molecular dynamics simulations of the photodissociated state of MbNO performed at 300 K using a fluctuating charge model for the nitric oxide (NO) ligand. After dissociation, NO is observed to remain mainly in the centre of the distal haem pocket, although some movement towards the primary docking site and the xenon-4 pocket can be seen. We calculate the NO infrared spectrum for the photodissociated ligand within the haem pocket and find a narrow peak in the range 1915-1922 cm(-1). The resulting blue shift of 1 to 8 cm(-1) compared to gas-phase NO is much smaller than the red shifts calculated and observed for carbon monoxide (CO) in Mb. A small splitting, due to NO in the xenon-4 pocket, is also observed. At lower temperatures, the spectra and conformational space explored by the ligand remain largely unchanged, but the electrostatic interactions with residue His64 become increasingly significant in determining the details of the ligand orientation within the distal haem pocket. The investigation of the effect of the L29F mutation reveals significant differences between the behaviour of NO and that of CO, and suggests a coupling between the ligand and the protein dynamics due to the different ligand dipole moments.

The reaction of a [(PHOX)Ir(COD)](+) complex (COD = 1,5-cyclooctadiene) with dihydrogen was studied by NMR spectroscopy (PHOX = chiral phosphinooxazoline ligand). A single [(PHOX)Ir(H)(2)(COD)](+) isomer was formed as the primary product at -40 degrees C in THF. Subsequent reaction with H(2) at -40 to 0 degrees C led to a mixture of two diastereomeric [(PHOX)Ir(H)(2)(solvent)(2)](+) complexes with concomitant loss of cyclooctane. The stereochemistry of the three hydride complexes could be assigned from the NMR data. The structures and energies of the observed hydride complexes and the possible stereoisomers were calculated using density functional theory. The substantial energy differences (up to 39 kcal/mol) between the various stereoisomers demonstrate the strong influence of the chiral ligand. The observed stereoselective formation of dihydride complexes can be explained by steric effects of the PHOX ligand combined with a strong electronic influence of the coordinating N and P atoms, favoring addition of a hydride trans to the Ir-N bond.

Tris(2-guanidinium-ethyl)amine (1) was prepared as a supramolecular receptor of hydrogen orthovanadate (HVO(4)(2-)) to mimic the active site of vanadium haloperoxidase (V-HPO). Both (1)H and (51)V NMR titration indicated 1:1 complex (5) formation between (1) with HVO(4)(2-) with a binding constant of 1.1 x 10(3) M(-1). Similar as V-HPO, a UV band at 307 nm was observed upon binding of HVO(4)(2-) to (1). According to DFT calculations UV transitions >300 nm observed for both the enzyme and its mimic are due to V-N interactions.

Molecular dynamics (MD) simulations (5-10ns in length) and normal mode analyses were performed for the monomer and dimer of native porcine insulin in aqueous solution; both starting structures were obtained from an insulin hexamer. Several simulations were done to confirm that the results obtained are meaningful. The insulin dimer is very stable during the simulation and remains very close to the starting X-ray structure; the RMS fluctuations calculated from the MD simulation agree with the experimental B-factors. Correlated motions were found within each of the two monomers; they can be explained by persistent non-bonded interactions and disulfide bridges. The correlated motions between residues B24 and B26 of the two monomers are due to non-bonded interactions between the side-chains and backbone atoms. For the isolated monomer in solution, the A chain and the helix of the B chain are found to be stable during 5ns and 10ns MD simulations. However, the N-terminal and the C-terminal parts of the B chain are very flexible. The C-terminal part of the B chain moves away from the X-ray conformation after 0.5-2.5ns and exposes the N-terminal residues of the A chain that are thought to be important for the binding of insulin to its receptor. Our results thus support the hypothesis that, when monomeric insulin is released from the hexamer (or the dimer in our study), the C-terminal end of the monomer (residues B25-B30) is rearranged to allow binding to the insulin receptor. The greater flexibility of the C-terminal part of the beta chain in the B24 (Phe-->Gly) mutant is in accord with the NMR results. The details of the backbone and side-chain motions are presented. The transition between the starting conformation and the more dynamic structure of the monomers is characterized by displacements of the backbone of Phe B25 and Tyr B26; of these, Phe B25 has been implicated in insulin activation.

We present potential energy surfaces for the He-N2H+ system adiabatically corrected for the zero-point motion along the intermolecular stretching vibrations v1 = 0 and v1 = 1. The potentials are extended to shorter He-N2H+ separations which makes them useful for scattering calculations. Close coupling calculations of the spinless S matrices for the rotational excitation of N2H+ by He are presented, and recoupling techniques to obtain collisional excitation cross sections between the N2H+ hyperfine levels are used. The propensity rules between hyperfine levels are investigated for the case where two nuclear spins are involved. It is found that the only well defined propensity rule is DeltaF = DeltaF1 = Deltaj and that calculations are required in order to obtain the relative intensities of the two-spin hyperfine cross sections., (Copyright 2004 American Institute of Physics)

The energetics and dynamics of double proton transfer (DPT) is investigated theoretically for the Watson-Crick conformation of the guanine-cytosine (GC) base pair. Using semiempirical density functional theory the isolated and DNA-embedded GC pair is considered. Differences in the energetics and dynamics of DPT thus addresses the question of how relevant studies of isolated base pairs are for the understanding of processes occurring in DNA. Two-dimensional potential energy surfaces involving the transferring hydrogen atoms and the proton donors and acceptors are presented for both systems. The DPT reaction is accompanied by a contraction of the distance between the two bases with virtually identical energetic barriers being 18.8 and 18.7 kcal/mol for the isolated and DNA-embedded system, respectively. However, the transition state for DPT in the DNA-embedded GC pair is offset by 0.1 A to larger N-H separation compared to the isolated GC pair. Using activated ab initio molecular dynamics, DPT is readily observed for the isolated base pair with a minimal amount of 21.4 kcal/mol of initial average kinetic energy along the DPT normal mode vector. On a time scale of approximately 100 fs DPT has occurred and the excess energy is redistributed. For the DNA-embedded GC pair considerably more kinetic energy is required (30.0 kcal/mol) for DPT and the process is completed within one hydrogen vibration. The relevance of studies of isolated base pairs and base pair analogs in regard of reactions or properties involving DNA is discussed., ((c) 2004 American Institute of Physics)

Possible insulin binding sites for D-glucose have been investigated theoretically by docking and molecular dynamics (MD) simulations. Two different docking programs for small molecules were used; Multiple Copy Simultaneous Search (MCSS) and Solvation Energy for Exhaustive Docking (SEED) programs. The configurations resulting from the MCSS search were evaluated with a scoring function developed to estimate the binding free energy. SEED calculations were performed using various values for the dielectric constant of the solute. It is found that scores emphasizing non-polar interactions gave a preferential binding site in agreement with that inferred from recent fluorescence and NMR NOESY experiments. The calculated binding affinity of -1.4 to -3.5 kcal/mol is within the measured range of -2.0 +/- 0.5 kcal/mol. The validity of the binding site is suggested by the dynamical stability of the bound glucose when examined with MD simulations with explicit solvent. Alternative binding sites were found in the simulations and their relative stabilities were estimated. The motions of the bound glucose during molecular dynamics simulations are correlated with the motions of the insulin side chains that are in contact with it and with larger scale insulin motions. These results raise the question of whether glucose binding to insulin could play a role in its activity. The results establish the complementarity of molecular dynamics simulations and normal mode analyses with the search for binding sites proposed with small molecule docking programs., (Copyright 2004 Wiley-Liss, Inc.)

Molecular dynamics simulations of the events after the photodissociation of CO in the myoglobin mutant L29F in which leucine is replaced by phenylalanine are reported. Using both classical and mixed quantum-classical molecular dynamics calculations, we observed the rapid motion of CO away from the distal heme pocket to other regions of the protein, in agreement with recent experimental results. The experimentally observed and calculated infrared spectra of CO after dissociation are also in good agreement. We compared the results with data from simulations of WT myoglobin. As the time resolution of experimental techniques is increased, theoretical methods and models can be validated at the atomic scale by direct comparison with experiment.

The dynamics of proton transfer along ammonia chains (chemical composition N(x)H(+)(3x+1), x=2, 4, and 6) in a constraining environment is investigated by ab initio molecular dynamics simulations. A carbon nanotube of defined length and diameter is used as an idealized constraining environment such that the ammonia chain is forced to maintain its quasilinear geometry. It is found that, although the energetics of proton transport shows considerable energetic barriers, proton translocation along the wire is possible at finite temperature for all chain lengths studied. The proton transport involves rotational reorientation of the proton-carrying ammonia molecule. High level ab initio calculations (MP2/aug-cc-pVTZ) yield barriers for internal rotation of 9.1 kcal/mol for NH(4) (+)-NH(3) and 11.7 kcal/mol for OH(3) (+)-OH(2), respectively. The infrared spectrum calculated from the dipole-dipole autocorrelation function shows distinct spectral features in the regions (2000-3000 cm(-1)) where the NHN proton transfer mode is expected to absorb. Assigning moderate opposite total charges between 0.002 and 0.2e to the carbon atoms at the end caps of the nanotube leads to a considerable speedup of the proton transfer., ((c) 2004 American Institute of Physics.)

Structural, energetic, and dynamical studies of Azotobacter vinelandii ferredoxin I are presented for native and mutant forms. The protein contains two iron-sulfur clusters, one of which ([3Fe-4S]) is believed to play a central role in the electron-coupled proton transfer. Different charge sets for the [3Fe-4S] cluster in its reduced and oxidized state are calculated with broken symmetry ab initio density functional theory methods and used in molecular dynamics (MD) simulations. The validity of the ab initio calculations is assessed by comparing partially optimized structures of the [3Fe-4S] clusters with x-ray structures. Possible proton transfer pathways between the protein and the iron-sulfur cluster are examined by both MD simulations and ab initio calculations. The MD simulations identify three main-chain hydrogen atoms--HN(13), HN(14), and HN(16)--that are within H-bonding distance of the [3Fe-4S] cluster throughout the MD simulations. They could thus play a role in the proton transfer from the protein to the iron-sulfur cluster. By contrast, the HD2(15) atom of the Asp-15 is seldom close enough to the [3Fe-4S] cluster to transfer a proton. Poisson-Boltzmann calculations indicate that there is a low, but nonzero probability, that Asp-15 is protonated at pH 7; this is a requirement for it to serve as a proton donor. Ab initio calculations with a fragment model for the protein find similar behavior for the transfer of a proton from the OH of the protonated side chain and the main-chain NH of Asp-15. The existence of a stable salt bridge between Asp-15 and Lys-84 in the D15E mutant, versus its absence in the wild-type, has been suggested as the cause of the difference in the rate of proton transfer. Extensive MD simulations were done to test this idea; the results do not support the proposal. The present findings, together with the available data, serve as the basis for an alternative proposal for the mechanism of the coupled electron-proton transfer reaction in ferredoxin I.

Molecular dynamics simulations of the photodissociated state of carbonmonoxy myoglobin (MbCO) are presented using a fluctuating charge model for CO. A new three-point charge model is fitted to high-level ab initio calculations of the dipole and quadrupole moment functions taken from the literature. The infrared spectrum of the CO molecule in the heme pocket is calculated using the dipole moment time autocorrelation function and shows good agreement with experiment. In particular, the new model reproduces the experimentally observed splitting of the CO absorption spectrum. The splitting of 3-7 cm(-1) (compared to the experimental value of 10 cm(-1)) can be directly attributed to the two possible orientations of CO within the docking site at the edge of the distal heme pocket (the B states), as previously suggested on the basis of experimental femtosecond time-resolved infrared studies. Further information on the time evolution of the position and orientation of the CO molecule is obtained and analyzed. The calculated difference in the free energy between the two possible orientations (Fe...CO and Fe...OC) is 0.3 kcal mol(-1) and agrees well with the experimentally estimated value of 0.29 kcal mol(-1). A comparison of the new fluctuating charge model with an established fixed charge model reveals some differences that may be critical for the correct prediction of the infrared spectrum and energy barriers. The photodissociation of CO from the myoglobin mutant L29F using the new model shows rapid escape of CO from the distal heme pocket, in good agreement with recent experimental data. The effect of the protein environment on the multipole moments of the CO ligand is investigated and taken into account in a refined model. Molecular dynamics simulations with this refined model are in agreement with the calculations based on the gas-phase model. However, it is demonstrated that even small changes in the electrostatics of CO alter the details of the dynamics.

Crown-capped iron(S-) porphyrins 1 x H2O and 2 x H2O and their corresponding Ba2+ complexes have been prepared as active site analogues of the resting state of cytochrome P450cam. cw-EPR studies and electronic structure calculations at the density functional theory (DFT) level of model systems suggest a functional role of the water cluster of P450cam.

The electron-coupled proton transfer reaction involving the [3Fe-4S] cluster in Ferredoxin I is studied by ab initio calculations and molecular dynamics simulations. The charge distributions of the [3Fe-4S] cluster are calculated with density functional theory (B3LYP) and used in the dynamics simulations. Structural differences between the oxidized and reduced clusters in the absence and presence of the protein are calculated to examine the hypothesis that an entatic state is involved in the reaction. The possible role of internal water molecules in the proton transfer process is explored. It is shown that water molecules are dynamically stable near the [3Fe-4S] cluster for tens of ps. Calculations for the native protein and a mutant D15N in which the Asp15 is replaced by a Glu15 are compared. It is found that water is less likely to escape from the region around the [3Fe-4S] cluster in the case of the D15N mutant than in the native protein. This finding could explain, in part, the lower proton transfer rate constant experimentally observed for the mutant if a water molecule were involved in transferring the proton from D15 to the [3Fe-4S] cluster.

The rebinding of NO to myoglobin after photolysis is studied using the 'reactive molecular dynamics' method. In this approach the energy of the system is evaluated on two potential energy surfaces that include the heme-ligand interactions which change between liganded and unliganded myoglobin. This makes it possible to take into account in a simple way, the high dimensionality of the transition seam connecting the reactant and product states. The dynamics of the dissociated NO molecules are examined, and the geometrical and energetic properties of the transition seam are studied. Analysis of the frequency of recrossing shows that the height of the effective rebinding barrier is dependent on the time after photodissociation. This effect is due mainly to protein relaxation and may contribute to the experimentally observed non-exponential rebinding rate of NO, as has been suggested previously.

Proton translocation along ammonia wires is investigated in 7-hydroxyquinoline.(NH(3))(n) clusters, both experimentally by laser spectroscopy and theoretically by Hartree-Fock and density functional (DFT) calculations. These clusters serve as realistic finite-size models for proton transfer along a chain of hydrogen-bonded solvent molecules. In the enol tautomer of 7-hydroxyquinoline (7-HQ), the OH group acts as a proton injection site into the (NH(3))(n)cluster. Proton translocation along a chain of three NH(3) molecules within the cluster can take place, followed by reprotonation of 7-HQ at the quinolinic N atom, forming the 7-ketoquinoline tautomer. Exoergic proton transfer from the OH group of 7-HQ to the closest NH(3) molecule within the cluster giving a zwitterion 7-HQ-.(NH(3))(6)H+ (denoted PT-A) occurs at a threshold cluster size of n = 6 in the DFT calculations and at n = 5 or 6 experimentally. Three further locally stable zwitterion clusters denoted PT-B, PT-B', and PT-C, the keto tautomer, and several transition structures along the proton translocation path were characterized theoretically. Grotthus-type proton-hopping mechanisms occur for three of the proton transfer steps, which have low barriers and are exoergic or weakly endoergic. The step with the highest barrier involves a complex proton transfer mechanism, involving structural reorganization and large-scale diffusive motions of the cluster.

The microwave spectra of N(2)-O(3) and N(2)-SO(2) have been recorded in the 6-18 GHz range using a pulsed-nozzle, Fourier transform microwave spectrometer. C-type transitions have been observed for both complexes which are slightly shifted by internal tunneling motions of the O(3) or SO(2) moieties. In addition, unshifted a-type transitions have been observed for N(2)-O(3). The nuclear hyperfine pattern is typical of equivalent nitrogen nuclei. Two sets of rotational and hyperfine constants are required to fit the symmetric and antisymmetric nuclear spin states, indicating that the equivalence arises from tunneling rotation of the nitrogen molecule. Internal tunneling motions along three tunneling pathways have been identified, although no information on the N(2) tunneling frequency is available from the spectra. From the N(2)-O(3) data the tunneling frequencies cannot be decorrelated from the rotational parameters; however, the O(3) tunneling frequency upper limit is estimated to be 2.0 MHz and the frequency of the concerted tunneling motion of both moieties is estimated to be about 8.9 MHz. For N(2)-SO(2), the SO(2) tunneling frequency is 11.5 kHz and the concerted frequency 173.9 kHz. Both complexes are roughly T shaped with the N(2) axis approximately perpendicular to the O(3) or SO(2) plane. In the equilibrium structures of both complexes, the a-c inertial plane is a plane of symmetry. The centers of mass separations are estimated from the rotational parameters to be 3.582 Å for N(2)-O(3) and 3.875 Å for N(2)-SO(2). The angle between the symmetry axes of the O(3) or SO(2) and the line joining their centers of mass have been calculated as 130.84 degrees (or 49.16 degrees ) and 119.71 degrees (or 60.29 degrees ), respectively. From the quadrupole analysis, the average angle between the N(2) axis and the a-inertial axis is 32.12 degrees for N(2)-O(3) and 27.81 degrees for N(2)-SO(2). Model electrostatic and ab initio calculations confirm these structures. Differences between the experimental and calculated structural parameters highlight the role of tunneling dynamics in these complexes. Copyright 2000 Academic Press.