Changes in the prostaglandin levels in alcohol toxicity: effect of curcumin and N-acetylcysteine

The present study was undertaken to evaluate the potential role of curcumin, the antioxidant principal from Curcuma longa Linn., and the sulphur-containing amino acid N-acetylcysteine against ethanol-induced changes in the levels of prostanoids. Biochemical assessment of liver damage was done by measuring the activities of serum enzymes (i.e., aspartate transaminase and alkaline phosphatase), which were significantly increased in rats fed ethanol, whereas the elevated levels of these enzymes were decreased after curcumin and N-acetylcysteine treatment to rats fed ethanol. We observed a significant increase in the levels of prostaglandins E(1), E(2), F(2alpha), and D(2) in liver, kidney, and brain. Administration of curcumin and N-acetylcysteine was shown to decrease the level of these prostanoids in the tissue studied.     

An all atom energy based computational protocol for predicting binding affinities of protein-ligand complexes

We report here a computationally fast protocol for predicting binding affinities of non-metallo protein-ligand complexes. The protocol builds in an all atom energy based empirical scoring function comprising electrostatics, van der Waals, hydrophobicity and loss of conformational entropy of protein side chains upon ligand binding. The method is designed to ensure transferability across diverse systems and has been validated on a heterogenous dataset of 161 complexes consisting of 55 unique protein targets. The scoring function trained on a dataset of 61 complexes yielded a correlation of r=0.92 for the predicted binding free energies against the experimental binding affinities. Model validation and parameter analysis studies ensure the predictive ability of the scoring function. When tested on the remaining 100 protein-ligand complexes a correlation of r=0.92 was recovered. The high correlation obtained underscores the potential applicability of the methodology in drug design endeavors. The scoring function has been web enabled at as binding affinity prediction of protein-ligand (BAPPL) server.     

The mu transpososome through a topological lens

Phage Mu is the most efficient transposable element known, its high efficiency being conferred by an enhancer DNA element. Transposition is the end result of a series of well choreographed steps that juxtapose the enhancer and the two Mu ends within a nucleoprotein complex called the 'transpososome.' The particular arrangement of DNA and protein components lends extraordinary stability to the transpososome and regulates the frequency, precision, directionality, and mechanism of transposition. The structure of the transpososome, therefore, holds the key to understanding all of these attributes, and ultimately to explaining the runaway genetic success of transposable elements throughout the biological world. This review focuses on the path of the DNA within the Mu transpososome, as uncovered by recent topological analyses. It discusses why Mu topology cannot be analyzed by standard methods, and how knowledge of the geometry of site alignment during Flp and Cre site-specific recombination was harnessed to design a new methodology called 'difference topology.' This methodology has also revealed the order and dynamics of association of the three interacting DNA sites, as well as the role of the enhancer in assembly of the Mu transpososome.     

p38 MAP Kinase and MAPKAP Kinases MK2/3 Cooperatively Phosphorylate Epithelial Keratins

The MAPK-activated protein kinases (MAPKAP kinases) MK2 and MK3 are directly activated via p38 MAPK phosphorylation, stabilize p38 by complex formation, and contribute to the stress response. The list of substrates of MK2/3 is increasing steadily. We applied a phosphoproteomics approach to compare protein phosphorylation in MK2/3-deficient cells rescued or not by ectopic expression of MK2. In addition to differences in phosphorylation of the known substrates of MK2, HSPB1 and Bag-2, we identified strong differences in phosphorylation of keratin 8 (K8). The phosphorylation of K8-Ser⁷³ is catalyzed directly by p38, which in turn shows MK2-dependent expression. Notably, analysis of small molecule p38 inhibitors on K8-Ser⁷³ phosphorylation also demonstrated reduced phosphorylations of keratins K18-Ser⁵² and K20-Ser¹³ but not of K8-Ser⁴³¹ or K18-Ser³³. Interestingly, K18-Ser⁵² and K20-Ser¹³ are not directly phosphorylated by p38 in vitro, but by MK2. Furthermore, anisomycin-stimulated phosphorylations of K20-Ser¹³ and K18-Ser⁵² are inhibited by small molecule inhibitors of both p38 and MK2. MK2 knockdown in HT29 cells leads to reduced K20-Ser¹³ phosphorylation, which further supports the notion that MK2 is responsible for K20 phosphorylation in vivo. Physiologic relevance of these findings was confirmed by differences of K20-Ser¹³ phosphorylation between the ileum of wild-type and MK2/3-deficient mice and by demonstrating p38- and MK2-dependent mucin secretion of HT29 cells. Therefore, MK2 and p38 MAPK function in concert to phosphorylate K8, K18, and K20 in intestinal epithelia.     

A role for siRNA in X-chromosome dosage compensation in Drosophila melanogaster

Sex-chromosome dosage compensation requires selective identification of X chromatin. How this occurs is not fully understood. We show that small interfering RNA (siRNA) mutations enhance the lethality of Drosophila males deficient in X recognition and partially rescue females that inappropriately dosage-compensate. Our findings are consistent with a role for siRNA in selective recognition of X chromatin.     

Maraviroc, a chemokine receptor-5 antagonist, fails to demonstrate efficacy in the treatment of patients with rheumatoid arthritis in a randomized, double-blind placebo-controlled trial

Introduction

The purpose of this study was to determine whether maraviroc, a human CC chemokine receptor 5 (CCR5) antagonist, is safe and effective in the treatment of active rheumatoid arthritis (RA) in patients on background methotrexate (MTX).

Methods

This phase IIa study comprised two distinct components: an open-label safety study of the pharmacokinetics (PK) of MTX in the presence of maraviroc, and a randomized, double-blind, placebo-controlled, proof-of-concept (POC) component. In the PK component, patients were randomized 1:1 to receive maraviroc 150 or 300 mg twice daily (BID) for four weeks. In the POC component, patients were randomized 2:1 to receive maraviroc 300 mg BID or placebo for 12 weeks. Patients were not eligible for inclusion in both components.

Results

Sixteen patients were treated in the safety/PK component. Maraviroc was well tolerated and there was no evidence of drug-drug interaction with MTX. One hundred ten patients were treated in the POC component. The study was terminated after the planned interim futility analysis due to lack of efficacy, at which time 59 patients (38 maraviroc; 21 placebo) had completed their week 12 visit. There was no significant difference in the number of ACR20 responders between the maraviroc (23.7%) and placebo (23.8%) groups (treatment difference -0.13%; 90% CI -20.45, 17.70; P = 0.504). The most common all-causality treatment-emergent adverse events in the maraviroc group were constipation (7.8%), nausea (5.2%), and fatigue (3.9%).

Conclusions

Maraviroc was generally well tolerated over 12 weeks; however, selective antagonism of CCR5 with maraviroc 300 mg BID failed to improve signs and symptoms in patients with active RA on background MTX.

Trial Registration

ClinicalTrials.gov: NCT00427934     

Halothiobacillus neapolitanus carboxysomes sequester heterologous and chimeric RubisCO species

Background

The carboxysome is a bacterial microcompartment that consists of a polyhedral protein shell filled with ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO), the enzyme that catalyzes the first step of CO₂ fixation via the Calvin-Benson-Bassham cycle.

Methodology/Principal Findings

To analyze the role of RubisCO in carboxysome biogenesis in vivo we have created a series of Halothiobacillus neapolitanus RubisCO mutants. We identified the large subunit of the enzyme as an important determinant for its sequestration into α-carboxysomes and found that the carboxysomes of H. neapolitanus readily incorporate chimeric and heterologous RubisCO species. Intriguingly, a mutant lacking carboxysomal RubisCO assembles empty carboxysome shells of apparently normal shape and composition.

Conclusions/Significance

These results indicate that carboxysome shell architecture is not determined by the enzyme they normally sequester. Our study provides, for the first time, clear evidence that carboxysome contents can be manipulated and suggests future nanotechnological applications that are based upon engineered protein microcompartments.     

Genetic regulation of amniotic fluid TNF-alpha and soluble TNF receptor concentrations affected by race and preterm birth

Racial disparity in spontaneous preterm birth (PTB) between African Americans and Caucasians in the US is unexplained, but is probably related to differences in amniotic fluid (AF) inflammatory cytokine profiles. Therefore, this study analyzed the association of 34 single nucleotide polymorphisms (SNPs) in TNF-α and its receptor genes (TNFR1 and TNFR2) with AF TNF-α and soluble TNF receptor (R1 and R2) concentrations in PTB. Samples consisted of African American and Caucasian cases (PTB), and controls (term birth) for which both cytokine, and maternal and fetal genotype data were available. Analyses were performed with genotype, case, and maker-status interaction in the model for log transformed cytokine concentrations. In Caucasians, two interactions between genotype and pregnancy outcome associated with cytokine concentrations, whereas 14 gene variants in African Americans showed interactions with pregnancy outcome, and 13 showed association with genetic markers. In conclusion, cytokine concentrations in African American preterm births can be partially explained by interactions between pregnancy outcome, SNPs and infection. This does not appear to be the case in Caucasians. These findings may be important in understanding disparity in rates of PTB between the two populations. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.