HIF-2α is indispensable for regulatory T cell function.

Hypoxia-inducible factor 1α (HIF-1α) and HIF-2α are master transcription factors that regulate cellular responses to hypoxia, but the exact function in regulatory T (Treg) cells is controversial. Here, we show that Treg cell development is normal in mice with Foxp3-specific knockout (KO) of HIF-1α or HIF-2α. However, HIF-2α-KO (but not HIF-1α-KO) Treg cells are functionally defective in suppressing effector T cell-induced colitis and inhibiting airway hypersensitivity.

Blocking IL-19 Signaling Ameliorates Allergen-Induced Airway Inflammation.

Asthma is a chronic inflammatory disease of the airway. Its major symptoms are reversible breathing problems causing airway narrowing and obstruction. IL-19 is a member of the IL-10 family cytokines.

Magnolol-mediated regulation of plasma triglyceride through affecting lipoprotein lipase activity in apolipoprotein A5 knock-in mice.

Hyperlipidemia is a risk factor of arteriosclerosis, stroke, and other coronary heart disease, which has been shown to correlate with single nucleotide polymorphisms of genes essential for lipid metabolism, such as lipoprotein lipase (LPL) and apolipoprotein A5 (APOA5). In this study, the effect of magnolol, the main active component extracted from Magnolia officinalis, on LPL activity was investigated. A dose-dependent up-regulation of LPL activity, possibly through increasing LPL mRNA transcription, was observed in mouse 3T3-L1 pre-adipocytes cultured in the presence of magnolol for 6 days.

Critical role of IL-6 in dendritic cell-induced allergic inflammation of asthma.

Unlabelled: Interleukin (IL)-6 plays important roles in autoimmunity and inflammation and is essential for T helper (Th) 2 and Th17 differentiation. However, whether it is involved in the development and function of dendritic cells (DCs) during allergen-induced airway inflammation and airway hyper-reactivity (AHR) remains undefined. In this study, Dermatophagoides pteronyssinus (Der p)-induced airway inflammation and AHR were studied in IL-6 knockout (KO) mice.

Computational study of the "DFG-flip" conformational transition in c-Abl and c-Src tyrosine kinases.

Protein tyrosine kinases are crucial to cellular signaling pathways regulating cell growth, proliferation, metabolism, differentiation, and migration. To maintain normal regulation of cellular signal transductions, the activities of tyrosine kinases are also highly regulated. The conformation of a three-residue motif Asp-Phe-Gly (DFG) near the N-terminus of the long "activation" loop covering the catalytic site is known to have a critical impact on the activity of c-Abl and c-Src tyrosine kinases.

Computational study of Gleevec and G6G reveals molecular determinants of kinase inhibitor selectivity.

Gleevec is a potent inhibitor of Abl tyrosine kinase but not of the highly homologous c-Src kinase. Because the ligand binds to an inactive form of the protein in which an Asp-Phe-Gly structural motif along the activation loop adopts a so-called DFG-out conformation, it was suggested that binding specificity was controlled by a "conformational selection" mechanism. In this context, the binding affinity displayed by the kinase inhibitor G6G poses an intriguing challenge.

An Overview of Electrostatic Free Energy Computations for Solutions and Proteins.

Free energy simulations for electrostatic and charging processes in complex molecular systems encounter specific difficulties owing to the long-range, 1/r Coulomb interaction. To calculate the solvation free energy of a simple ion, it is essential to take into account the polarization of nearby solvent but also the electrostatic potential drop across the liquid-gas boundary, however distant. The latter does not exist in a simulation model based on periodic boundary conditions because there is no physical boundary to the system.

Computational analysis of the binding specificity of Gleevec to Abl, c-Kit, Lck, and c-Src tyrosine kinases.

Gleevec, a well-known cancer therapeutic agent, is an effective inhibitor of several tyrosine kinases, including Abl and c-Kit, but displays less potency to inhibit closely homologous tyrosine kinases, such as Lck and c-Src. Because many structural features of the binding site are highly conserved in these homologous kinases, the molecular determinants responsible for the binding specificity of Gleevec remain poorly understood. To address this issue, free energy perturbation molecular dynamics (FEP/MD) simulations with explicit solvent was used to compute the binding affinity of Gleevec to Abl, c-Kit, Lck, and c-Src.

Explaining why Gleevec is a specific and potent inhibitor of Abl kinase.

Tyrosine kinases present attractive drug targets for specific types of cancers. Gleevec, a well-known therapeutic agent against chronic myelogenous leukemia, is an effective inhibitor of Abl tyrosine kinase. However, Gleevec fails to inhibit closely homologous tyrosine kinases, such as c-Src.

Functional importance of apolipoprotein A5 185G in the activation of lipoprotein lipase.

Background: Apolipoprotein A5 (APOA5) over-expression enhances lipolysis of triglyceride (TG) through stimulation of lipoprotein lipase (LPL) activity; however, an APOA5 G185C variant was found associated with hypertriglyceridemia. The aim of this study was, therefore, to explore the importance of APOA5 185GG in the activation of LPL.

Methods: A fragment containing mature human APOA5 cDNA was obtained by RT-PCR and subcloned into pET-15b vector.

Lactobacillus gasseri suppresses Th17 pro-inflammatory response and attenuates allergen-induced airway inflammation in a mouse model of allergic asthma.

Probiotics are normal inhabitants of the gastrointestinal tract of man and are widely considered to exert a number of beneficial effects in many diseases. But the mechanism by which they modulate the immune system is poorly understood. The present study was planned to explore the anti-allergic effect of Lactobacillus gasseri on a mouse model of allergic asthma.

Molecular dynamics simulations of the intramolecular proton transfer and carbanion stabilization in the pyridoxal 5'-phosphate dependent enzymes L-dopa decarboxylase and alanine racemase.

Molecular dynamics simulations using a combined quantum mechanical and molecular mechanical (QM/MM) potential have been carried out to investigate the internal proton transfer equilibrium of the external aldimine species in l-dopa decarboxylase, and carbanion stabilization by the enzyme cofactor in the active site of alanine racemase. Solvent effects lower the free energy of the O-protonated PLP tautomer both in aqueous solution and in the active site, resulting a free energy difference of about -1 kcal/mol relative to the N-protonated Schiff base in the enzyme. The external aldimine provides the dominant contribution to lowering the free energy barrier for the spontaneous decarboxylation of l-dopa in water, by a remarkable 16 kcal/mol, while the enzyme l-dopa decarboxylase further lowers the barrier by 8 kcal/mol.

Kinetic isotope effects of L-Dopa decarboxylase.

A mixed centroid path integral and free energy perturbation method (PI-FEP/UM) has been used to investigate the primary carbon and secondary hydrogen kinetic isotope effects (KIEs) in the amino acid decarboxylation of L-Dopa catalyzed by the enzyme L-Dopa decarboxylase (DDC) along with the corresponding uncatalyzed reaction in water. DDC is a pyridoxal 5'-phosphate (PLP) dependent enzyme. The cofactor undergoes an internal proton transfer between the zwitterionic protonated Schiff base configuration and the neutral hydroxyimine tautomer.

The opsin shift and mechanism of spectral tuning in rhodopsin.

Molecular dynamics simulations and combined quantum mechanical and molecular mechanical calculations have been performed to investigate the mechanism of the opsin shift and spectral tuning in rhodopsin. A red shift of -980 cm(-1) was estimated in the transfer of the chromophore from methanol solution environment to the protonated Schiff base (PSB)-binding site of the opsin. The conformational change from a 6-s-cis-all-trans configuration in solution to the 6-s-cis-11-cis conformer contributes additional -200 cm(-1), and the remaining effects were attributed to dispersion interactions with the aromatic residues in the binding site.

A Non-Orthogonal Block-Localized Effective Hamiltonian Approach for Chemical and Enzymatic Reactions.

The effective Hamiltonian-molecular orbital and valence bond (EH-MOVB) method based on non-orthogonal block-localized fragment orbitals has been implemented into the program CHARMM for molecular dynamics simulations of chemical and enzymatic reactions, making use of semiempirical quantum mechanical models. Building upon ab initio MOVB theory, we make use of two parameters in the EH-MOVB method to fit the barrier height and the relative energy between the reactant and product state for a given chemical reaction to be in agreement with experiment or high-level ab initio or density functional results. Consequently, the EH-MOVB method provides a highly accurate and computationally efficient QM/MM model for dynamics simulation of chemical reactions in solution.

Randomized placebo-controlled trial of lactobacillus on asthmatic children with allergic rhinitis.

Previous studies have suggested that probiotic administration may have therapeutic and/or preventive effects on atopic dermatitis in infants; however, its role in allergic airway diseases remains controversial. To determine whether daily supplementation with specific Lactobacillus gasseri A5 for 8 weeks can improve the clinical symptoms and immunoregulatory changes in school children suffering from asthma and allergic rhinitis (AR). We conducted a randomized, double-blind, placebo-controlled study on school children (age, 6-12 years) with asthma and AR.

Internal proton transfer in the external pyridoxal 5'-phosphate Schiff base in dopa decarboxylase.

Combined quantum mechanical and molecular mechanical (QM/MM) simulations of dopa decarboxylase have been carried out to elucidate the factors that contribute to the tautomeric equilibrium of the intramolecular proton transfer in the external PLP-L-dopa Schiff base. The presence of a carboxylate anion on the alpha-carbon of the Schiff base stabilizes the zwitterions and shifts the equilibrium in favor of the oxoenamine tautomer (protonated Schiff base). Moreover, protonation of the PLP pyridine nitrogen further drives the equilibrium toward the oxoenamine direction.

Explicit polarization (X-Pol) potential using ab initio molecular orbital theory and density functional theory.

The explicit polarization (X-Pol) method has been examined using ab initio molecular orbital theory and density functional theory. The X-Pol potential was designed to provide a novel theoretical framework for developing next-generation force fields for biomolecular simulations. Importantly, the X-Pol potential is a general method, which can be employed with any level of electronic structure theory.

Comparison of nitric oxide-induced oxidation of recombinant oxyhemoglobin subunits using a competition experiment.

A low reaction rate with nitric oxide (NO) is one of the important characteristics of hemoglobin (Hb)-based oxygen carriers. The reaction rate between oxyHb and NO is usually measured by stopped-flow spectrophotometry. However, the reported rates vary due to the difficulty of accurately determining the NO concentration and the limit of the instrument dead time.

Common sequence variant in lipoprotein lipase gene conferring triglyceride response to fibrate treatment.

Aims: Little is known about whether the variations in the lipoprotein lipase (LPL) and apolipoprotein gene cluster (APOA1/C3/A5) confer appreciable triglyceride lowering after fibrate treatment among patients with hypertriglyceridemia.

Materials & Methods: We investigated whether variations in these genes confer a triglyceride lowering response after fibrate treatment among 145 patients with hypertriglyceridemia receiving 6 months of fibrate treatment.

Results: Overall triglycerides decreased from 746.

Hemoglobin conjugated with a Band 3 N-terminus derived peptide as an oxygen carrier.

A peptide composed of 9 amino acids, 7 residues from N-terminus of human erythrocytic Band 3 protein (AcMEELQDD) followed by cysteine and glutamic acids, was conjugated to hemoglobin (Hb) serving as an allosteric effector for oxygen release. The activated polyethylene glycol (PEG), maleimide-PEG-N-hydroxysuccinimidyl, was used to crosslink Hb with the peptide. The putative conjugation site on Hb for effective enhancement of oxygen release was characterized as Lys-beta95 by liquid chromatography-tandem mass spectrometry.

Hybrid quantum and classical methods for computing kinetic isotope effects of chemical reactions in solutions and in enzymes.

A method for incorporating quantum mechanics into enzyme kinetics modeling is presented. Three aspects are emphasized: 1) combined quantum mechanical and molecular mechanical methods are used to represent the potential energy surface for modeling bond forming and breaking processes, 2) instantaneous normal mode analyses are used to incorporate quantum vibrational free energies to the classical potential of mean force, and 3) multidimensional tunneling methods are used to estimate quantum effects on the reaction coordinate motion. Centroid path integral simulations are described to make quantum corrections to the classical potential of mean force.

APOA1/C3/A5 haplotype and risk of hypertriglyceridemia in Taiwanese.

Background: Apolipoprotein A5 gene (APOA5) has been shown to modulate plasma triglyceride concentrations. We investigated 2 distinct APOA1/C3/A5 haplotypes roles for hypertriglyceridemia.

Methods: We recruited 308 cases of hypertriglyceridemia and 281 normal controls from a hospital.

Differential effects of the Zn-His-Bkb vs Zn-His-[Asp/Glu] triad on Zn-core stability and reactivity.

The most common partner of the Zn-bound His is the Asp/Glu carboxylate side chain in catalytic Zn sites and the backbone (Bkb) carbonyl group in structural Zn sites. To elucidate the factors governing the selection of the second-shell partner of the Zn-bound His in structural/catalytic Zn sites, systematic studies using density functional theory and continuum dielectric calculations were performed to determine the relative contributions of the second-shell Bkb carbonyl and the Asp/Glu carboxylate to the Zn-core stability and reactivity. The results show that the contributions of the second-shell Bkb carbonyl and Asp/Glu carboxylate to the Zn-core stability depend mainly on the solvent accessibility of the Zn-site and the composition of the Zn-core.

Factors governing the protonation state of Zn-bound histidine in proteins: a DFT/CDM study.

We have performed systematic theoretical studies to elucidate the factors governing the His protonation/deprotonation state in Zn-binding sites, especially those containing the ubiquitous Zn-His-Asp/Glu triad. Specifically, we have addressed the following three questions: (1) How does the transfer of the Zn-bound His imidazole proton to the second-shell Asp/Glu carboxylate oxygen depend on the composition of the other first-shell ligands and the solvent accessibility of the metal-binding site? (2) Can any second-shell ligand with a proton acceptor group such as the backbone carbonyl oxygen also act as a proton acceptor? (3) What is the effect of the Asp/Glu in the Zn-His-Asp/Glu triad on the Zn-bound water protonation state? To address these questions, we used a combination of quantum mechanical and continuum dielectric methods to compute the free energies for deprotonating a Zn-bound imidazole/water in various Zn complexes. The calculations show that whether the Zn-bound His is protonated or deprotonated depends on (1) the solvent accessibility of the metal-binding site, and (2) the Lewis acid ability of Zn, which is indirectly determined by both the first- and the second-shell Zn ligands.