Tjerk P. Straatsma

Differential tapasin dependence of MHC class I molecules correlates with conformational changes upon peptide dissociation: a molecular dynamics simulation study.

Efficiency of peptide loading to MHC class I molecules in the endoplasmic reticulum is allele specific and can involve interaction with tapasin and other proteins. Allele HLA-B 4,402 depends on tapasin whereas HLA-B 4,405 (Tyr116 instead of Asp in B 4,402) can efficiently load peptides without tapasin. Both alleles adopt very similar structures in the presence of the same peptide. Molecular dynamics simulations on peptide termini dissociation from the alpha(1)/alpha(2) binding domains were used to characterize structural and free energy changes. The magnitude of the calculated free energy change and the shape of the free energy curve vs. distance for induced peptide C terminus dissociation differed for B 4,405 compared to B 4,402. Structural changes during C terminus dissociation occurred mainly in the first segment of the alpha(2)-helix that flanks the peptide C terminus binding region (F pocket) and contacts residue 116. This segment is also close to the proposed tapasin contact region. For B 4402, a stable shift towards an altered open F pocket structure deviating significantly from the bound form was observed. In contrast, B 4405 showed only a transient opening of the F pocket followed by relaxation towards a structure close to the bound (receptive) form upon C terminus dissociation. The greater tendency for a peptide-receptive conformation in the absence of peptide combined with more long-range interactions with the peptide C terminus facilitates peptide binding to B 4405 and correlates with the tapasin-independence of this allele. A possible role of tapasin in case of HLA-B 4402 and other tapasin-dependent alleles could be the stabilization of a peptide-receptive class I conformation.

Characterization of the outer membrane protein OprF of Pseudomonas aeruginosa in a lipopolysaccharide membrane by computer simulation.

The N‐terminal domain of outer membrane protein OprF of Pseudomonas aeruginosa forms a membrane spanning eight‐stranded antiparallel β‐barrel domain that folds into a membrane channel with low conductance. The structure of this protein has been modeled after the crystal structure of the homologous protein OmpA of Escherichia coli. A number of molecular dynamics simulations have been carried out for the homology modeled structure of OprF in an explicit molecular model for the rough lipopolysaccharide (LPS) outer membrane of P. aeruginosa. The structural stability of the outer membrane model as a result of the strong electrostatic interactions compared with simple lipid bilayers is restricting both the conformational flexibility and the lateral diffusion of the porin in the membrane. Constricting side‐chain interactions within the pore are similar to those found in reported simulations of the protein in a solvated lipid bilayer membrane. Because of the strong interactions between the loop regions of OprF and functional groups in the saccharide core of the LPS, the entrance to the channel from the extracellular space is widened compared with the lipid bilayer simulations in which the loops are extruding in the solvent. The specific electrostatic signature of the LPS membrane, which results in a net intrinsic dipole across the membrane, is found to be altered by the presence of OprF, resulting in a small electrically positive patch at the position of the channel. Proteins 2009.

Assessment of the convergence of molecular dynamics simulations of lipopolysaccharide membranes

The outer membrane of Gram-negative bacteria is composed of a phospholipid inner leaflet and a lipopolysaccharide (LPS) outer leaflet. The chemical structure of LPS results in an asymmetric character of outer membranes that has been shown to play an important role in the electrical properties of porins, low permeability and intrinsic antibiotic resistance of Gram-negative bacteria. Atomistic molecular dynamics simulations of two different configurations of the outer membrane of Pseudomonas aeruginosa under periodic boundary conditions were carried out in order to (1) validate model-derived properties against the available experimental data, (2) identify the properties whose dynamics can be sampled on nanosecond timescales, and (3) evaluate the dependence of the convergence of structural and dynamical properties on the initial configuration of the system, within the chosen force field and simulation conditions. Because the relaxation times associated with the motions of individual LPS monomers in outer membranes are very long, the two initial configurations do not converge to a common ensemble of configuration on the nanosecond time scale. However, a number of properties of the outer membrane that will significantly impact the structural and internal dynamics of transmembrane proteins, most notably the electrostatic potential and molecular density, do converge within the simulated time scale. For these properties, a good agreement with the available experimental data was found. Such a molecular model, capable of accounting for the high asymmetry and low fluidity characteristics of outer membranes provides a more appropriate environment for atomistic simulations of outer membrane proteins.

Multiplet splittings and other properties from density functional theory: an assessment in iron–porphyrin systems

In transition metal compounds with spin states close in energy, the magnitude and sign of the energy splitting calculated with density functional theory depends strongly on the functional used. Therefore we must turn to additional criteria to assess the level of accuracy and reliability of predictions based on this level of theory. We report optimized geometries, total energies, and Mössbauer quadrupole splitting values for low-spin and high-spin, ferric and ferrous model hemes using a variety of gradient-corrected and hybrid functionals. In one model, the iron–porphyrin is axially ligated by two strong-field imidazole ligands [FeP(Im)2] and has a low-spin ground state. In the other model complex the axial ligands are two weak-field, water molecules [FeP(H2O)2], and have a high-spin ground state. Among all the functionals used (UHF, B3LYP, B3LYP*, BLYP, half-and-half, LSDA), the B3LYP hybrid functional most consistently reproduced the experimental geometry, Mössbauer, and spin state data for the two model hemes. Simply gradient-corrected functionals exhibit strong biases towards low spin states, while Hartree–Fock favours strongly high spin states. These findings suggest that for systems with similar characteristics of several accessible electronic spin configurations, it is imperative to include properties other than just the energy in the assessment of the DFT predictions.

Interaction between the CBM of Cel9A from Thermobifida fusca and Cellulose Fibers

Molecular docking and molecular dynamics (MD) simulations were used to investigate the binding of a cellodextrin chain in a crystal‐like conformation to the carbohydrate‐binding module (CBM) of Cel9A from Thermobifida fusca. The fiber was found to bind to the CBM in a single and well‐defined configuration in‐line with the catalytic cleft, supporting the hypothesis that this CBM plays a role in the catalysis by feeding the catalytic domain (CD) with a polysaccharide chain. The results also expand the current known list of residues involved in the binding. The polysaccharide‐protein attachment is shown to be mediated by five amine/amide‐containing residues. E478 and E559 were found not to interact directly with the sugar chain; instead they seem to be responsible to stabilize the binding motif via hydrogen bonds. Copyright

Force Field Development and Molecular Dynamics of [NiFe] Hydrogenase

Classical molecular force-field parameters describing the structure and motion of metal clusters in [NiFe] hydrogenase enzymes can be used to compare the dynamics and thermodynamics of [NiFe] under different oxidation, protonation, and ligation circumstances. Using density functional theory (DFT) calculations of small model clusters representative of the active site and the proximal, medial, and distal Fe/S metal centers and their attached protein side chains, we have calculated classical force-field parameters for [NiFe] in reduced and oxidized states, including internal coordinates, force constants, and atom-centered charges. Derived force constants revealed that cysteinate ligands bound to the metal ions are more flexible in the Ni-B active site, which has a bridging hydroxide ligand, than in the Ni-C active site, which has a bridging hydride. Ten nanosecond all-atom, explicit-solvent MD simulations of [NiFe] hydrogenase in oxidized and reduced catalytic states established the stability of the derived force-field parameters in terms of Cα and metal cluster fluctuations. Average active site structures from the protein MD simulations are consistent with [NiFe] structures from the Protein Data Bank, suggesting that the derived force-field parameters are transferrable to other hydrogenases beyond the structure used for testing. A comparison of experimental H2-production rates demonstrated a relationship between cysteinate side chain rotation and activity, justifying the use of a fully dynamic model of [NiFe] metal cluster motion.

Engineering an ultra-stable affinity reagent based on Top7

Antibodies are widely used for diagnostic and therapeutic applications because of their sensitive and specific recognition of a wide range of targets; however, their application is limited by their structural complexity. More demanding applications require greater stability than can be achieved by immunoglobulin-based reagents. Highly stable, protein-based affinity reagents are being investigated for this role with the goal of identifying a suitable scaffold that can attain specificity and sensitivity similar to that of antibodies while performing under conditions where antibodies fail. We have engineered Top7--a highly stable, computationally designed protein--to specifically bind human CD4 by inserting a peptide sequence derived from a CD4-specific antibody. Molecular dynamics simulations were used to evaluate the structural effect of the peptide insertion at a specific site within Top7 and suggest that this Top7 variant retains conformational stability over 100 degrees C. This engineered protein specifically binds CD4 and, consistent with simulations, is extremely resistant to thermal and chemical denaturation--retaining its secondary structure up to at least 95 degrees C and requiring 6 M guanidine to completely unfold. This CD4-specific protein demonstrates the functionality of Top7 as a viable scaffold for use as a general affinity reagent which could serve as a robust and inexpensive alternative to antibodies.

A Multiprotocol Communication Support for the Global Address Space Programming Model on the IBM SP

The paper describes an efficient communication support for the global address space programming model on the IBM SP, a commercial example of the SMP (symmetric multi-processor) clusters. Our approach integrates shared memory with active messages, threads and remote memory copy between nodes. The shared memory operations offer substantial performance improvement over LAPI, IBM one-sided communication library, within an SMP node. Based on the experiments with the SPLASH-2 LU benchmark and a molecular dynamics simulation, our multiprotocol support for the global address space is found to improve performance and scalability of applications. This approach could also be used in optimizing the MPI-2 one-sided communication on the SMP clusters.

Molecular basis for microbial adhesion to geochemical surfaces: computer simulation of Pseudomonas aeruginosa adhesion to goethite.

The adhesion of Pseudomonas aeruginosa to the goethite mineral is investigated using classical molecular simulation. A fragment model for goethite has been integrated into a fully atomistic membrane model. Properties for the resulting system are evaluated for a 1.5-ns simulation in the isothermal-isobaric ensemble. The response of the membrane to the presence of the mineral is investigated. Radial distribution functions are used to present an average picture of the hydrogen bonding. Orientational vectors, assigned to the saccharide groups, reveal the extent of the minerals perturbations on the membrane. Significant structural changes were observed for the outermost saccharide groups, several of which rotate to form hydrogen bonds with the mineral surface. The structure of the inner core, and the corresponding integrity of the membrane, is maintained. The mineral surface dehydrates slightly in the presence of the membrane as saccharide hydroxyl groups compete with water molecules for hydrogen-bonding sites on its surface.

Influence of the B-band O-antigen chain in the structure and electrostatics of the lipopolysaccharide membrane of Pseudomonas aeruginosa

Lipopolysaccharides (LPSs) form the major constituent of the outer membrane of Gram-negative bacteria, and are believed to play a key role in processes that govern microbial metal binding, surface adhesion, and microbe-mediated oxidation/reduction reactions. It is also a major causative agent of nosocomial illness, eliciting both chronic and acute infections in burn, immunocompromised, and cystic fibrosis. Phenotypic variation in the relative expression of A- and B-band in the LPS of Pseudomonas aeruginosa seems to alter its overall surface characteristics influencing adhesion and favoring survival. Classical molecular dynamics simulations of A-B+ LPS membrane model of P. aeruginosa were carried out in explicit solvent for 12+ ns. The B-band presents a remarkable flexibility remaining fully solvated and does not interact with the sugar units from the LPS core surface residues, in agreement with atomic force microscopy experiments. Comparison with previous simulations of the rough LPS membrane suggests that the presence of the B-band promotes membrane expansion. In addition, this O-antigen chain dramatically alters the electrostatic potential and surface charge of the LPS membrane. This is illustrated by the resulting electrostatic surface potential. These results are compared to previous simulations of the rough LPS and a model hypothesis is proposed to explain the increased ability of B-band expressing microorganisms to adhere to cell surfaces and the necessity of these organisms to loose the O side chain for the development of acute infections.