The first description of a (1--> 6)-beta-D-glucan in prokaryotes: (1 --> 6)-beta-D-glucan is a common component of actinobacillus suis and is the basis for a serotyping system

The chemical and antigenic properties of the cell-surface lipopolysaccharides (LPSs) and capsular polysaccharides (CPSs) of seven representative strains of Actinobacillus suis from healthy and diseased pigs were investigated. Four strains produced a linear (1 --> 6)-beta-D-glucan homopolymer, beta-D-Glcp-(1-[ --> 6)-beta-D-Glcp-(1-]n -->, as a LPS-O-chain (O1) and as a CPS (K1). Polyclonal antisera prepared against a (1 --> 6)-beta-D-glucan-containing strain showed a positive reaction against both LPSs and CPSs derived from the above strains (designated serotype O1/K1). One strain carried the (1 --> 6)-beta-D-glucan solely as a LPS-O-chain (serotype O1) and two strains did not express the (1 --> 6)-beta-D-glucan, but, instead, produced a different O-chain (designated serotype 02); these three strains expressed their own characteristic CPSs. (1 --> 6)-beta-D-Glucan structures are common cell wall components of yeast, fungi and lichens, but, to our knowledge, this is the first time a (1 --> 6)-beta-D-glucan has been described in a prokaryotic organism. Conformational and nuclear magnetic resonance analyses showed that the beta-D-Glcp-(1 --> 6)-beta-D-Glcp linkage was flexible and two distinct glycosidic conformers are described. Cross-reactive antibodies to the A. suis (1 --> 6)-beta-D-glucan could be detected in sera from a variety of species and in sera from specific pathogen free pigs. This cross-reactivity may arise from immuno-stimulation of organisms present in the surrounding environment that contain (1 --> 6)-beta-D-glucan, which may also explain the high incidence of false positive results in previous serological tests for A. suis. In addition, these (1 --> 6)-beta-D-glucan background antibodies may be protective against A. suis infection. The characterization herein of (1 --> 6)-beta-D-glucan is the foundation for the development of a serotyping system for A. suis.     

Stator structure and subunit composition of the V(1)/V(0) Na(+)-ATPase of the thermophilic bacterium Caloramator fervidus

The V-type Na(+)-ATPase of the thermophilic, anaerobic bacterium Caloramator fervidus was purified to homogeneity. The subunit compositions of the catalytic V(1) and membrane-embedded V(0) parts were determined and the structure of the enzyme complex was studied by electron microscopy. The V(1) headpiece consists of seven subunits present in one to three copies, and the V(0) part of two subunits in a ratio of 5:2. An analysis of over 7500 single particle images obtained by electron microscopy of the purified V(1)V(0) enzyme complex revealed that the stalk region, connecting the V(1) and V(0) parts, contains two peripheral stalks in addition to a central stalk. One of the two is connected to the V(0) part, while the other is connected to the first via a bar-like structure that is positioned just above V(0), parallel with the plane of the membrane. In projection, this bar seems to contact the central stalk. The data show that the stator structure that prevents rotation of the static part of V(0) relative to V(1) in the rotary catalysis mechanism of energy coupling in ATPases/ATPsynthases is more complex than previously thought.     

Sequential and structural homology between intracellular pathogenesis-related proteins and a group of latex proteins

The intracellular pathogenesis-related proteins have been identified in a broad range of flowering plants. Some display quite different patterns of expression, in many cases unrelated to the pathogenic response. Nevertheless, these proteins are all very similar and in most cases share more than 35% sequence identity. In this report we investigate the significance of a rather weak similarity between the intracellular pathogenesis-related (IPR or PR-10) proteins and a group of proteins identified in the latex of opium poppy and in Arabidopsis, among others. A sequence analysis held together with the recently published three-dimensional structure of Bet v 1, an IPR protein from birch pollen, strongly suggests sequential and structural homology between the two protein families.     

Crystal Structures of DNA-Whirly Complexes and Their Role in Arabidopsis Organelle Genome Repair

DNA double-strand breaks are highly detrimental to all organisms and need to be quickly and accurately repaired. Although several proteins are known to maintain plastid and mitochondrial genome stability in plants, little is known about the mechanisms of DNA repair in these organelles and the roles of specific proteins. Here, using ciprofloxacin as a DNA damaging agent specific to the organelles, we show that plastids and mitochondria can repair DNA double-strand breaks through an error-prone pathway similar to the microhomology-mediated break-induced replication observed in humans, yeast, and bacteria. This pathway is negatively regulated by the single-stranded DNA (ssDNA) binding proteins from the Whirly family, thus indicating that these proteins could contribute to the accurate repair of plant organelle genomes. To understand the role of Whirly proteins in this process, we solved the crystal structures of several Whirly-DNA complexes. These reveal a nonsequence-specific ssDNA binding mechanism in which DNA is stabilized between domains of adjacent subunits and rendered unavailable for duplex formation and/or protein interactions. Our results suggest a model in which the binding of Whirly proteins to ssDNA would favor accurate repair of DNA double-strand breaks over an error-prone microhomology-mediated break-induced replication repair pathway.     

The impact of herpes zoster and postherpetic neuralgia on health-related quality of life: a prospective study

Background

Vaccination against herpes zoster is being considered in many countries. We conducted a multicentre prospective study to describe the impact of herpes zoster and postherpetic neuralgia on health-related quality of life.

Methods

From October 2005 to July 2006, 261 outpatients aged 50 years or older with herpes zoster were recruited from the clinical practices of 83 physicians within 14 days after rash onset. The Zoster Brief Pain Inventory was used to measure severity of pain and interference with activities of daily living because of pain. The EuroQol EQ-5D assessment tool was used to measure quality of life. These outcomes were assessed at recruitment and on days 7, 14, 21, 30, 60, 90, 120, 150 and 180 following recruitment.

Results

Acute herpes zoster interfered in all health domains, especially sleep (64% of participants), enjoyment of life (58%) and general activities (53%). The median duration of pain was 32.5 days. The median duration of interference with activities of daily living because of pain varied between 27 and 30 days. Overall, 24% of the participants had postherpetic neuralgia (pain for more than 90 days after rash onset). Anxiety and depression, enjoyment of life, mood and sleep were most frequently affected during the postherpetic neuralgia period. The mean EQ-5D score was 0.59 at enrolment and remained at 0.67 at all follow-up points among participants who reported clinically significant pain.

Interpretation

These data support the need for preventive strategies and additional early intervention to reduce the burden of herpes zoster and postherpetic neuralgia.Herpes zoster, which is characterized by dermatomal pain and vesicular rash,^1,2 results from reactivation of the varicella-zoster virus.^3,4 The average lifetime risk of herpes zoster in developed countries is estimated to be about 30%^5–7 and increases with increasing life expectancy. The most common complication of herpes zoster, and one of the most challenging to treat, is postherpetic neuralgia, a painful condition often defined as pain persisting for more than 90 days after rash onset.⁸ According to this definition, postherpetic neuralgia is estimated to occur in 8%–27% of people with herpes zoster overall.^9–14 The risk of postherpetic neuralgia increases markedly with age.¹⁵The Shingles Prevention Study, a randomized double-blind placebo-controlled trial, showed that a live-attenuated varicella-zoster virus vaccine was safe and effective in preventing herpes zoster and postherpetic neuralgia among people 60 years of age and older.¹³ Given these promising results, policy-makers and clinicians are being asked to make recommendations regarding the use and funding of the herpes zoster vaccine. To do this, evidence on the burden of herpes zoster from the patient’s perspective is required. The only data available on the impact of herpes zoster on health-related quality of life comes from two short-term studies.^16,17 Clinical reports and cross-sectional surveys^18–20 have also suggested that postherpetic neuralgia can profoundly impair quality of life. However, no study followed a cohort of patients with newly diagnosed herpes zoster for a sufficient period to assess postherpetic neuralgia and describe the associated impact on quality of life.We undertook a multicentre prospective study to describe the impact of herpes zoster and postherpetic neuralgia on health-related quality of life.     

Genetic analysis of 103 candidate genes for coronary artery disease and associated phenotypes in a founder population reveals a new association between endothelin-1 and high-density lipoprotein cholesterol

Coronary artery disease (CAD) is a major health concern in both developed and developing countries. With a heritability estimated at ~50%, there is a strong rationale to better define the genetic contribution to CAD. This project involves the analysis of 884 individuals from 142 families (with average sibships of 5.7) as well as 558 case and control subjects from the Saguenay Lac St-Jean region of northeastern Quebec, with the use of 1,536 single-nucleotide polymorphisms (SNPs) in 103 candidate genes for CAD. By use of clusters of SNPs to generate multiallelic haplotypes at candidate loci for segregation studies within families, suggestive linkage for high-density lipoprotein (HDL) cholesterol is observed on chromosome 1p36.22. Furthermore, several associations that remain significant after Bonferroni correction are observed with lipoprotein-related traits as well as plasma concentrations of adiponectin. Of note, HDL cholesterol levels are associated with an amino acid substitution (lysine/asparagine) at codon 198 (rs5370) of endothelin-1 (EDN1) in a sex-specific manner, as well as with a SNP (rs2292318) located 7.7 kb upstream of lecithin cholesterol acyl-transferase (LCAT). Whereas the other observed associations are described in the current literature, these two are new. Using an independent validation sample of 806 individuals, we confirm the EDN1 association (P<.005), whereas the LCAT association was nonsignificant (P=.12).     

Identification and biochemical characterization of two novel UDP-2,3-diacetamido-2,3-dideoxy-alpha-D-glucuronic acid 2-epimerases from respiratory pathogens

For a long period lactate was considered as a dead-end product of glycolysis in many cells and its accumulation correlated with acidosis and cellular and tissue damage. At present, the role of lactate in several physiological processes has been investigated based on its properties as an energy source, a signalling molecule and as essential for tissue repair. It is noteworthy that lactate accumulation alters glycolytic flux independently from medium acidification, thereby this compound can regulate glucose metabolism within cells. PFK (6-phosphofructo-1-kinase) is the key regulatory glycolytic enzyme which is regulated by diverse molecules and signals. PFK activity is directly correlated with cellular glucose consumption. The present study shows the property of lactate to down-regulate PFK activity in a specific manner which is not dependent on acidification of the medium. Lactate reduces the affinity of the enzyme for its substrates, ATP and fructose 6-phosphate, as well as reducing the affinity for ATP at its allosteric inhibitory site at the enzyme. Moreover, we demonstrated that lactate inhibits PFK favouring the dissociation of enzyme active tetramers into less active dimers. This effect can be prevented by tetramer-stabilizing conditions such as the presence of fructose 2,6-bisphosphate, the binding of PFK to f-actin and phosphorylation of the enzyme by protein kinase A. In conclusion, our results support evidence that lactate regulates the glycolytic flux through modulating PFK due to its effects on the enzyme quaternary structure.     

Development of specific inhibitors for heparin-binding proteins based on the cobra cardiotoxin structure: an effective synthetic strategy for rationally modified heparin-like disaccharides and a trisaccharide

Recently, a new heparin disaccharide-binding site on the convex side of cobra cardiotoxin (CTX) was identified by NMR spectroscopy and molecular modeling. To further characterize this site two heparin-like disaccharides were synthesized for binding studies with CTX, and a trisaccharide was synthesized for testing the sequence of the disaccharide binding to CTX. Thus six differentially protected monosaccharide building blocks (three l-iduronic acids and three d-glucosamines) were prepared. These include a l-iduronic acid elongation building block namely methyl 2-O-acetyl-4-O-levulinoyl-3-O-pivaloyl-alpha-l-idopyranosyluronate trichloroacetimidate for which a single-crystal X-ray structure was determined to have M(r)=576.79, a=9.3098(11)A alpha=90 degrees , b=10.3967(12)A beta=90 degrees , c=28.026(3)A gamma=90 degrees , V=2712.7(6)A(3), P2(1)2(1)2(1), Z=4, mu=0.71073A, and R=0.0378 for 7586 observed reflections. It shows that the molecular structure of the donor is in the (1)C(4) conformation with significant 1,3-diaxial interactions between O-1 and O-3 as well as O-2 and O-4. The disaccharides and trisaccharide vary in the degree and position of O- and N-sulfation. The pivaloyl group was used as permanent protecting group of hydroxyl. The levulinoyl group was used as the temporary protecting group to protect the hydroxyl for elongation.