Eosinophils, but not eosinophil peroxidase or major basic protein, are important for host protection in experimental Brugia pahangi infection

The attenuation of eosinophilia by the administration of monoclonal antibodies to CCR3 consistently correlates with impairment in worm elimination following primary intraperitoneal Brugia pahangi infections in mice. Host protection was unimpaired in mice deficient in eosinophil peroxidase (EPO) or major basic protein 1 (MBP-1), suggesting that eosinophils are essential in host protection but that neither EPO nor MBP-1 alone is.     

Aqueous urea solution destabilizes Abeta(16-22) oligomers

We use long multiple trajectories generated by molecular dynamics simulations to probe the stability of oligomers of Abeta(16-22) (KLVFFAE) peptides in aqueous urea solution. High concentration of urea promotes the formation of beta-strand structures in Abeta(16-22) monomers, whereas in water they adopt largely compact random coil structures. The tripeptide system, which forms stable antiparallel beta-sheet structure in water, is destabilized in urea solution. The enhancement of beta-strand content in the monomers and the disruption of oligomeric structure occur largely by direct interaction of urea with the peptide backbone. Our simulations suggest that the oligomer unbinding dynamics is determined by two opposing effects, namely, by the increased propensity of monomers to form beta-strands and the rapid disruption of the oligomers. The qualitative conclusions are affirmed by using two urea models. Because the proposed destabilization mechanism depends largely on hydrogen bond formation between urea and the peptide backbone, we predict that high urea concentration will destabilize oligomers of other amyloidogenic peptides as well.     

Dynamics of unbinding of cell adhesion molecules: transition from catch to slip bonds

The unbinding dynamics of complexes involving cell-adhesion molecules depends on the specific ligands. Atomic force microscopy measurements have shown that for the specific P-selectin-P-selectin glycoprotein ligand (sPSGL-1) the average bond lifetime t initially increases (catch bonds) at low (< or =10 pN) constant force, f, and decreases when f > 10 pN (slip bonds). In contrast, for the complex with G1 anti-P-selectin monoclonal antibody t monotonically decreases with f. To quantitatively map the energy landscape of such complexes we use a model that considers the possibility of redistribution of population from one force-free state to another force-stabilized bound state. The excellent agreement between theory and experiments allows us to extract energy landscape parameters by fitting the calculated curves to the lifetime measurements for both sPSGL-1 and G1. Surprisingly, the unbinding transition state for P-selectin-G1 complex is close (0.32 nm) to the bound state, implying that the interaction is brittle, i.e., once deformed, the complex fractures. In contrast, the unbinding transition state of the P-selectin-sPSGL-1 complex is far (approximately 1.5 nm) from the bound state, indicative of a compliant structure. Constant f energy landscape parameters are used to compute the distributions of unbinding times and unbinding forces as a function of the loading rate, rf. For a given rf, unbinding of sPSGL-1 occurs over a broader range of f with the most probable f being an order of magnitude less than for G1. The theory for cell adhesion complexes can be used to predict the outcomes of unbinding of other protein-protein complexes.     

Ribosome exit tunnel can entropically stabilize alpha-helices

Several experiments have suggested that newly synthesized polypeptide chains can adopt helical structures deep within the ribosome exit tunnel. We hypothesize that confinement in the roughly cylindrical tunnel can entropically stabilize alpha-helices. The hypothesis is validated by using theory and simulations of coarse-grained off-lattice models. The model helix, which is unstable in the bulk, is stabilized in a cylindrical cavity provided the diameter (D) of the cylinder exceeds a critical value D*. When D < D* both the helical content and the helix-coil transition temperature (T(f)) decrease abruptly. Surprisingly, we find that the stability of the alpha-helix depends on the number (N) of amino acid residues. Entropic stabilization, as measured by changes in T(f), increases nonlinearly as N increases. The simulation results are in quantitative agreement with a standard helix-coil theory that takes into account entropy cost of confining a polypeptide chain in a cylinder. The results of this work are in qualitative accord with most of the findings of a recent experiment in which N-dependent ribosome-induced helix stabilization of transmembrane sequences was measured by fluorescence resonance energy transfer.     

Mechanisms and kinetics of beta-hairpin formation

Thermodynamics and kinetics of off-lattice models with side chains for the beta-hairpin fragment of immunoglobulin-binding protein and its variants are reported. For all properties (except refolding time tau(F)) there are no qualitative differences between the full model and the Go version. The validity of the models is established by comparison of the calculated native structure with the Protein Data Bank coordinates and by reproducing the experimental results for the degree of cooperativity and tau(F). For the full model tau(F) approximately 2 micros at the folding temperature (experimental value is 6 micros); the Go model folds 50 times faster. Upon refolding, structural changes take place over three time scales. On the collapse time scale compact structures with intact hydrophobic cluster form. Subsequently, hydrogen bonds form, predominantly originating from the turn by a kinetic zipping mechanism. The assembly of the hairpin is complete when most of the interstrand contacts (the rate-limiting step) is formed. The dominant transition state structure (located by using cluster analysis) is compact and structured. We predict that when hydrophobic cluster is moved to the loop tau(F) marginally increases, whereas moving the hydrophobic cluster closer to the termini results in significant decrease in tau(F) relative to wild type. The mechanism of hairpin formation is predicted to depend on turn stiffness.     

Minimum energy compact structures in force-quench polyubiquitin folding are domain swapped

Single molecule experiments that initiate folding using mechanical force are uniquely suited to reveal the nature of populated states in the folding process. Using a strategy proposed on theoretical grounds, which calls for repeated cycling of force from high to low values using force pulses, it was demonstrated in atomic force spectroscopy (AFM) experiments that an ensemble of minimum energy compact structures (MECS) are sampled during the folding of polyubiquitin. The structures in the ensemble are mechanically resistant to a lesser extent than the native state. Remarkably, forced unfolding of the populated intermediates reveals a broad distribution of extensions including steps up to 30 nm and beyond. We show using molecular simulations that favorable interdomain interactions leading to domain swapping between adjacent ubiquitin modules results in the formation of the ensemble of MECS, whose unfolding leads to an unusually broad distribution of steps. We obtained the domain-swapped structures using coarse-grained ubiquitin dimer models by exchanging native interactions between two monomeric ubiquitin molecules. Brownian dynamics force unfolding of the proposed domain-swapped structures, with mechanical stability that is approximately 100-fold lower than the native state, gives rise to a distribution of extensions from 2 to 30 nm. Our results, which are in quantitative agreement with AFM experiments, suggest that domain swapping may be a general mechanism in the assembly of multi-sub-unit proteins.     

IL-13Ralpha2 and IL-10 coordinately suppress airway inflammation, airway-hyperreactivity, and fibrosis in mice

Development of persistent Th2 responses in asthma and chronic helminth infections are a major health concern. IL-10 has been identified as a critical regulator of Th2 immunity, but mechanisms for controlling Th2 effector function remain unclear. IL-10 also has paradoxical effects on Th2-associated pathology, with IL-10 deficiency resulting in increased Th2-driven inflammation but also reduced airway hyperreactivity (AHR), mucus hypersecretion, and fibrosis. We demonstrate that increased IL-13 receptor alpha 2 (IL-13Ralpha2) expression is responsible for the reduced AHR, mucus production, and fibrosis in BALB/c IL-10(-/-) mice. Using models of allergic asthma and chronic helminth infection, we demonstrate that IL-10 and IL-13Ralpha2 coordinately suppress Th2-mediated inflammation and pathology, respectively. Although IL-10 was identified as the dominant antiinflammatory mediator, studies with double IL-10/IL-13Ralpha2-deficient mice illustrate an indispensable role for IL-13Ralpha2 in the suppression of AHR, mucus production, and fibrosis. Thus, IL-10 and IL-13Ralpha2 are both required to control chronic Th2-driven pathological responses.     

Dynamics of allosteric transitions in GroEL

The chaperonin GroEL-GroES, a machine that helps proteins to fold, cycles through a number of allosteric states, the T state, with high affinity for substrate proteins, the ATP-bound R state, and the R" (GroEL-ADP-GroES) complex. Here, we use a self-organized polymer model for the GroEL allosteric states and a general structure-based technique to simulate the dynamics of allosteric transitions in two subunits of GroEL and the heptamer. The T --> R transition, in which the apical domains undergo counterclockwise motion, is mediated by a multiple salt-bridge switch mechanism, in which a series of salt-bridges break and form. The initial event in the R -->R" transition, during which GroEL rotates clockwise, involves a spectacular outside-in movement of helices K and L that results in K80-D359 salt-bridge formation. In both the transitions there is considerable heterogeneity in the transition pathways. The transition state ensembles (TSEs) connecting the T, R, and R" states are broad with the TSE for the T --> R transition being more plastic than the R --> R" TSE.