Constant pH Molecular Dynamics with Proton Tautomerism

The current article describes a new two-dimensional λ-dynamics method to include proton tautomerism in continuous constant pH molecular dynamics (CPHMD) simulations. The two-dimensional λ-dynamics framework is used to devise a tautomeric state titration model for the CPHMD simulations involving carboxyl and histidine residues. Combined with the GBSW implicit solvent model, the new method is tested on titration simulations of blocked histidine and aspartic acid as well as two benchmark proteins, turkey ovomucoid third domain (OMTKY3) and ribonuclease A (RNase A). A detailed analysis of the errors inherent to the CPHMD methodology as well as those due to the underlying solvation model is given. The average absolute error for the computed pK(a) values in OMTKY3 is 1.0 pK unit. In RNase A the average absolute errors for the carboxyl and histidine residues are 1.6 and 0.6 pK units, respectively. In contrast to the previous work, the new model predicts the correct sign for all the pK(a) shifts, but one, in the benchmark proteins. The predictions of the tautomeric states of His(12) and His(48) and the conformational states of His(48) and His(119) are in agreement with experiment. Based on the simulations of OMTKY3 and RNase A, the current work has demonstrated the capability of the CPHMD technique in revealing pH-coupled conformational dynamics of protein side chains

Systemic effects of oral tolerance on inflammation: mobilization of lymphocytes and bone marrow eosinopoiesis

Oral tolerance is a T-cell mediated phenomenon defined by inhibition of immune responsiveness to a protein previously contacted by the oral route. Oral tolerance may prevent autoimmune and allergic diseases that involve the recruitment and/or activation of different cell types including mast cells, neutrophils, eosinophils, monocytes and lymphocytes. The mechanisms by which oral tolerance avoids these immunological disorders are still controversial. Herein we used a murine model of ovalbumin (OVA)-induced peritonitis to investigate the effect of oral tolerance on allergic inflammation. Frequency of leucocyte subpopulations was evaluated by global and differential cell counts in peritoneal lavage fluid, peripheral blood, and bone marrow. Changes on lymphocyte subsets and adhesion molecules expression by these cells were analysed by flow cytometry. As compared with OVA-immune mice, intraperitoneal challenge of tolerant animals with OVA resulted in a significantly milder peritonitis, mostly affecting neutrophils and eosinophils; a concomitant reduction in total white blood cell counts was also observed, mainly because of lower neutrophil and eosinophil counts. Eosinophils, but not neutrophils, were also reduced in the bone-marrow of OVA-challenged tolerant mice. No changes occurred in total peritoneal lymphocyte counts in OVA-tolerant mice, however, there was a significant decrease in CD3(+) CD8(+) T cells and an increase in B cells (CD45R(+)) in these animals as compared to immune OVA-challenged animals. Altered expression of CD18 and CD54, respectively, in blood and peritoneal lymphocytes was also noted. These results suggest that, in addition to local specific effects, oral tolerance has systemic effects on the mobilization of leucocytes and bone-marrow eosinopoiesis

Characterizing RNA Dynamics at Atomic Resolution Using Solution-state NMR Spectroscopy

Many recently discovered non-coding RNAs do not fold into a single native conformation, but rather, sample many different conformations along their free energy landscape to carry out their biological function. Unprecedented insights into the RNA dynamic structure landscape are provided by solution-state NMR techniques that measure the structural, kinetic, and thermodynamic characteristics of motions spanning picosecond to second timescales at atomic resolution. From these studies a basic description of the RNA dynamic structure landscape is emerging, bringing new insights into how RNA structures change to carry out their function as well as applications in RNA-targeted drug discovery and RNA bioengineering

Nickel oral hyposensitization in patients with systemic nickel allergy syndrome

Background: This is the fi rst randomized, double-blind, placebo- controlled trial (EUDRACT No. 2009-013923-43) evaluating nickel oral hyposensitizing treatment (NiOHT) in patients with \u201c systemic nickel allergy syndrome \u201d (SNAS), characterized by Niallergic contact dermatitis and systemic reactions after eating Ni-rich food. Methods: Adults with positive Ni-patch test, who reported symptoms suggesting SNAS, which improved after Ni-poor diet, and were positive to Ni-oral challenge were eligible. Patients were randomly assigned to three treatments (1.5 g, 0.3 g, or 30 ng Ni/week) or placebo for a year, with progressive reintroduction of Ni-rich foods form the 5 th month. Out of 141 patients randomized, 113 completed the trial. Endpoints were effi cacy and tolerability of treatment. Results: During Ni-rich food re-introduction, the 1.5 g Ni/week group had a mean VAS score signifi cantly higher than placebo (p 0.044), with signifi cant improvement of gastrointestinal symptoms (p 0.016;) and signifi cantly fewer rescue medications. Cutaneous manifestations also improved but without reaching statistical signifi cance. After the treatment, oral challenge with higher Ni doses than at baseline were needed to cause symptoms to fl are-up in signifi cantly more patients given 1.5 g Ni/week than placebo (p 0.05). Patients reported no side-eff ects. Conclusions: NiOHT is eff ective in SNAS, in particular on gastrointestinal manifestations, with trend toward improvement of cutaneous symptoms