Disparate impact of oxidative host defenses determines the fate of Salmonella during systemic infection in mice.

Reactive oxygen and nitrogen species function in host defense via mechanisms that remain controversial. Pathogens might encounter varying levels of these species, but bulk measurements cannot resolve such heterogeneity. We used single-cell approaches to determine the impact of oxidative and nitrosative stresses on individual Salmonella during early infection in mouse spleen. Salmonella encounter and respond to both stresses, but the levels and impact vary widely. Neutrophils and inflammatory monocytes kill Salmonella by generating overwhelming oxidative stress through NADPH oxidase and myeloperoxidase. This controls Salmonella within inflammatory lesions but does not prevent their spread to more permissive resident red pulp macrophages, which generate only sublethal oxidative bursts. Regional host expression of inducible nitric oxide synthase exposes some Salmonella to nitrosative stress, triggering effective local Salmonella detoxification through nitric oxide denitrosylase. Thus, reactive oxygen and nitrogen species influence dramatically different outcomes of disparate Salmonella-host cell encounters, which together determine overall disease progression.

Factors controlling material deposition in the CVD of nickel sulfides, selenides or phosphides from dichalcogenoimidodiphosphinato complexes: deposition, spectroscopic and computational studies.

The series of nickel dichalcogenoimidodiphosphinates [Ni{(i)Pr(2)P(X1)NP(X2)(i)Pr(2)}(2)]: X1 = S, X2 = Se (1), X1 = X2 = S (2), and X1 = X2 = Se (3) have been successfully used as single-source precursors (SSPs) to deposit thin films of nickel sulfide, selenide or phosphide; the material deposited depended on both temperature and method used for the deposition. Aerosol-assisted (AA) chemical vapour deposition (CVD) and low-pressure (LP) CVD were used. The as-deposited films were characterised by X-ray diffraction (XRD), scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX) and X-ray photoelectron spectroscopy (XPS). A variety of phases including: Ni(2)P, Ni(0.85)Se and NiS(1.03) were deposited under different conditions. The mechanism of decomposition to the phosphide, selenide, or sulfide was studied by pyrolysis gas chromatography mass spectrometry (Py-GC-MS) and modelled by density functional theory (DFT).

The effects of perfluorination on carbohydrate-pi interactions: computational studies of the interaction of benzene and hexafluorobenzene with fucose and cyclodextrin.

The effect of benzene fluorination on C-H...pi interactions is studied using a number of computational methods applied to a range of intermolecular complexes. High level wavefunction methods (CCSD(T)) predict a slightly greater interaction energy for complexes of benzene with methane or fucose, compared to corresponding complexes involving hexafluorobenzene. A number of more approximate treatments, DFT with the M06-2X functional, PM3-D* and MM methods, give interaction energies within 1 kcal mol(-1) of the high level values, and also correctly predict that the interaction energy is slightly greater for benzene compared to hexafluorobenzene. However, the DFT-D model used here predicts that the interaction energy is slightly greater for hexafluorobenzene. Molecular dynamics simulations, employing the GLYCAM-06 force field, validated here, are used to model the complexes of benzene and hexafluorobenzene with beta-cyclodextrin in aqueous solution. We predict the binding free energies of the complexes to be within 0.5 kcal mol(-1), and suggest that the different chemical shifts of the H5 protons observed in the two complexes arise from their slightly different structures, rather than from different binding energies.

Inhibitors of PIM-1 kinase: a computational analysis of the binding free energies of a range of imidazo [1,2-b] pyridazines.

The binding of a selection of competitive imidazo [1,2-b] pyridazine inhibitors of PIM-1 kinase with nanomolar activity has been analyzed using computational methods. Molecular dynamics simulations using umbrella sampling to determine a potential of mean force have been used to accurately predict the relative free energies of binding of these inhibitors, from -4.3 to -9.5 kcal mol(-1), in excellent agreement with the trends observed in previous experimental assays. The relative activity of the inhibitors could not be accounted for by any single effect or interaction within the active site and could only be fully reproduced when the overall free energies were considered, including important contributions from interactions outside the hinge region and using explicit solvent in the active site. The potential of mean force for the displacement of the glycine-rich phosphate binding loop (P-loop) has also been estimated and shown to be an important feature in the binding of these ligands.

How are the ready and unready states of nickel-iron hydrogenase activated by H2? A density functional theory study.

We have explored possible mechanisms for the formation of the catalytically active Ni(a)-S state of the enzyme, nickel iron hydrogenase, from the Ni*(r) (ready) or Ni*(u) (unready) state, by reaction with H(2), using density functional theory calculations with the BP86 functional in conjunction with a DZVP basis set. We find that for the reaction of the ready state, which is taken to have an -OH bridge, the rate determining step is the cleavage of H(2) at the Ni(3+) centre with a barrier of approximately 15 kcal mol(-1). We take the unready state to have a -OOH bridge, and find that reaction with H(2) to form the Ni(r)-S state can proceed by two possible routes. One such path has a number of steps involving electron transfer, which is consistent with experiment, as is the calculated barrier of approximately 19 kcal mol(-1). The alternative pathway, with a lower barrier, may not be rate determining. Overall, our predictions give barriers in line with experiment, and allow details of the mechanism to be explored which are inaccessible from experiment.

Structure and redox properties of the protein, rubredoxin, and its ligand and metal mutants studied by electronic structure calculation.

The changes in the structural and electronic properties accompanying metal ionization of the iron-containing protein, rubredoxin, and of some ligand and metal mutants, have been explored using density functional theory (DFT) calculations of the metal atom coordinated to the four immediate residues. Both isolated and embedded cluster studies have been carried out, the latter using the hybrid quantum mechanics/molecular mechanics (QM/MM) approach. The replacement of a cysteine by a serine residue has a considerable effect on both the electronic and geometric structure of the core, which can be qualitatively understood on the basis of the isolated cluster studies. The modulation of these properties caused by the protein environment is quite accurately described by the QM/MM calculations. The predicted core geometries are in good accord with both X-ray and EXAFS data, and the changes in the redox potentials are predicted, at least semiquantitatively, by considering only the core part of the protein.

Theoretical analysis of peptidyl alpha-ketoheterocyclic inhibitors of human neutrophil elastase: Insight into the mechanism of inhibition and the application of QM/MM calculations in structure-based drug design.

It has been suggested from QSAR data (P. D. Edwards, D. J. Wolanin, D.A. Andisik and M. W. Davis, J. Med. Chem., 1995, 38, 76) that the inhibition of elastase by peptidyl alpha-ketoheterocyclic inhibitors can occur in two ways, the less potent inhibitors forming a non-bonded Michaelis complex and the more potent set a covalently bonded enzyme-substrate intermediate. We report QM/MM studies of both binding and reactivity that confirm these findings, showing that the activity of the least potent set of inhibitors correlates with the calculated binding energy, and that of the more potent set correlates with the stability of the intermediate. These calculations show that QM/MM methods can be successfully employed to understand complicated structure-activity relationships and might be employed in the design and assessment of new inhibitors.

Prediction of the potency of inhibitors of adenosine deaminase by QM/MM calculations.

QM/MM calculations show that the potency of a range of inhibitors of adenosine deaminase correlates with the relative stability of the reaction intermediate at the active site, rather than with the inhibitor binding energy.