The Application of a Laboratory Induction Furnace to the Selective Smelting of a Montana Chromite Concentrate

The purpose of this experimental work was to determine with the utilization of a laboratory sized induction furnace a method whereby a high-iron Montana chromite concentrate could be successfully smelted to yield a product suitable for the subsequent production of standard ferrochrome

Characterization of a dual function macrocyclase enables design and use of efficient macrocyclization substrates

H.L. is funded by the George & Stella Lee Scholarship and Criticat EPSRC. This project was also funded by the European Research Council project 339367 NCB-TNT and by the BBSRC (K015508/1). JHN is 1000 talent scholar of the Chinese Academy of Sciences at the University of Sichuan.Peptide macrocycles are promising therapeutic molecules because they are protease resistant, structurally rigid, membrane permeable and capable of modulating protein-protein interactions. Here, we report the characterization of the dual function macrocyclase-peptidase enzyme involved in the biosynthesis of the highly toxic Amanitin toxin family of macrocycles. The enzyme first removes 10 residues from the N-terminus of a 35-residue substrate. Conformational trapping of the amino acid peptide forces the enzyme to release this intermediate rather than proceed to macrocyclization. The enzyme rebinds the 25 amino acid peptide in a different conformation and catalyzes macrocyclization of the N-terminal 8 residues. Structures of the enzyme bound to both substrates and biophysical analysis characterize the different binding modes rationalizing the mechanism. Using these insights simpler substrates with only five C-terminal residues were designed, allowing the enzyme to be more effectively exploited in biotechnology.Publisher PDFPeer reviewe

Erioflorin stabilizes the tumor suppressor Pdcd4 by inhibiting its interaction with the E3-ligase β-TrCP1

Loss of the tumor suppressor Pdcd4 was reported for various tumor entities and proposed as a prognostic marker in tumorigenesis. We previously characterized decreased Pdcd4 protein stability in response to mitogenic stimuli, which resulted from p70S6K1-dependent protein phosphorylation, β-TrCP1-mediated ubiquitination, and proteasomal destruction. Following high-throughput screening of natural product extract libraries using a luciferase-based reporter assay to monitor phosphorylation-dependent proteasomal degradation of the tumor suppressor Pdcd4, we succeeded in showing that a crude extract from Eriophyllum lanatum stabilized Pdcd4 from TPA-induced degradation. Erioflorin was identified as the active component and inhibited not only degradation of the Pdcd4-luciferase-based reporter but also of endogenous Pdcd4 at low micromolar concentrations. Mechanistically, erioflorin interfered with the interaction between the E3-ubiquitin ligase β-TrCP1 and Pdcd4 in cell culture and in in vitro binding assays, consequently decreasing ubiquitination and degradation of Pdcd4. Interestingly, while erioflorin stabilized additional β-TrCP-targets (such as IκBα and β-catenin), it did not prevent the degradation of targets of other E3-ubiquitin ligases such as p21 (a Skp2-target) and HIF-1α (a pVHL-target), implying selectivity for β-TrCP. Moreover, erioflorin inhibited the tumor-associated activity of known Pdcd4- and IκBα-regulated αtranscription factors, that is, AP-1 and NF-κB, altered cell cycle progression and suppressed proliferation of various cancer cell lines. Our studies succeeded in identifying erioflorin as a novel Pdcd4 stabilizer that inhibits the interaction of Pdcd4 with the E3-ubiquitin ligase β-TrCP1. Inhibition of E3-ligase/target-protein interactions may offer the possibility to target degradation of specific proteins only as compared to general proteasome inhibition

Desferrioxamine biosynthesis : diverse hydroxamate assembly by substrate-tolerant acyl transferase DesC

Hydroxamate groups play key roles in the biological function of diverse natural products. Important examples include trichostatin A, which inhibits histone deacetylases via coordination of the active site zinc(II) ion with a hydroxamate group, and the desferrioxamines, which use three hydroxamate groups to chelate ferric iron. Desferrioxamine biosynthesis in Streptomyces species involves the DesD-catalysed condensation of various N-acylated derivatives of N-hydroxycadaverine with two molecules of N-succinyl-N-hydroxycadaverine to form a range of linear and macrocyclic tris-hydroxamates. However, the mechanism for assembly of the various N-acyl-N-hydroxycadaverine substrates of DesD from N-hydroxycadaverine has until now been unclear. Here we show that the desC gene of Streptomyces coelicolor encodes the acyl transferase responsible for this process. DesC catalyses the N-acylation of N-hydroxycadaverine with acetyl, succinyl and myristoyl-CoA, accounting for the diverse array of desferrioxamines produced by S. coelicolor. The X-ray crystal structure of DesE, the ferrioxamine lipoprotein receptor, in complex with ferrioxamine B (which is derived from two units of N-succinyl-N-hydroxycadaverine and one of N-acetyl-N-hydroxycadaverine) was also determined. This shows that the acetyl group of ferrioxamine B is solvent exposed, suggesting that the corresponding acyl group in longer chain congeners can protrude from the binding pocket, providing insights into their likely functio

Oligosaccharide and Glycoprotein Microarrays as Tools in HIV Glycobiology Glycan-Dependent gp120/Protein Interactions

AbstractDefining HIV envelope glycoprotein interactions with host factors or binding partners advances our understanding of the infectious process and provides a basis for the design of vaccines and agents that interfere with HIV entry. Here we employ carbohydrate and glycoprotein microarrays to analyze glycan-dependent gp120-protein interactions. In concert with new linking chemistries and synthetic methods, the carbohydrate arrays combine the advantages of microarray technology with the flexibility and precision afforded by organic synthesis. With these microarrays, we individually and competitively determined the binding profiles of five gp120 binding proteins, established the carbohydrate structural requirements for these interactions, and identified a potential strategy for HIV vaccine development

Periplasmic depolymerase provides insight into ABC transporter-dependent secretion of bacterial capsular polysaccharides

This work was supported in part by grants from the Canadian Institutes of Health Research (FDN_148364) (to C.W.). S.D.L. is a recipient of a Natural Science and Engineering Research Council Alexander Graham Bell Canada Graduate Scholarship and Michael Smith Foreign Study Supplement. C.W. is a Canada Research Chair. J.H.N. is a Wellcome Trust Investigator (100209/Z/12/Z).Capsules are surface layers of hydrated capsular polysaccharides (CPSs) produced by many bacteria. The human pathogen Salmonella enterica serovar Typhi produces "Vi antigen" CPS, which contributes to virulence. In a conserved strategy used by bacteria with diverse CPS structures, translocation of Vi antigen to the cell surface is driven by an ATP-binding cassette (ABC) transporter. These transporters are engaged in heterooligomeric complexes proposed to form an enclosed translocation conduit to the cell surface, allowing the transporter to power the entire process. We identified Vi antigen biosynthesis genetic loci in genera of the Burkholderiales, which are paradoxically distinguished from S. Typhi by encoding VexL, a predicted pectate lyase homolog. Biochemical analyses demonstrated that VexL is an unusual metal-independent endolyase with an acidic pH optimum that is specific for Oacetylated Vi antigen. A 1.22-Å crystal structure of the VexL-Vi antigen complex revealed features which distinguish common secreted catabolic pectate lyases from periplasmic VexL, which participates in cell-surface assembly. VexL possesses a right-handed parallel beta-superhelix, of which one face forms an electropositive glycan-binding groove with an extensive hydrogen bonding network that includes Vi antigen acetyl groups and confers substrate specificity. VexL provided a probe to interrogate conserved features of the ABC transporter-dependent export model. When introduced into S. Typhi, VexL localized to the periplasm and degraded Vi antigen. In contrast, a cytosolic derivative had no effect unless export was disrupted. These data provide evidence that CPS assembled in ABC transporter-dependent systems is actually exposed to the periplasm during envelope translocation.Publisher PDFPeer reviewe

799-2 Left Ventricular (LV) and Myocyte Electrophysiology with the Development of Dilated Cardiomyopathy (DCM); Effects of Angiotensin II Receptor (AT1 AT-II) Blockade

Ventricular arrhythmias are a significant cause of morbidity and mortality with DCM, and AT1 AT-II receptor activation has been implicated to play a role in arrhythmogenesis. However, the effects of AT1, AT-II receptor activation on changes in LV function and myocyte electrophysiology during the progression of DCM remain unexplored. Accordingly, this study measured weekly changes in LV function (ejection fraction, LVEF; peak systolic wall stress, LVWS) and surface electrocardiography (R-R interval, QRS duration, QTc interval), and myocyte action potentials (resting membrane, RM; upstroke velocity, Vmax; duration at 90% repolarization, APD90) at terminal study in 3 groups of dogs (n=6/group): DCM, chronic pace (216 bpm, 4 weeks); DCM/AT-BLOCK, chronic pace and treatment with a specific non-peptide AT1 AT-II antagonist (SR 47436 (BMS 186295); 30mg/kg BID); and control (CON). All measurements were made with the pacemaker deactivated.LVEF (%)LVWS (g/cm2)R-R (ms).QRS (ms).QTc (ms)Week 2:CON68.7±3.2133±14646±9958.4±1.3291±13DCM40.9±4.1*184±16*519±4060.7±1.9316±9DCM/AT-Block44.1±3.7*138±10+540±566.32±1.2*325±9Week4:CON73.1±2.4127±10629±4557.6±1.4314±9DCM35.2±3.5*223±16*505±41*62.0±1.9313±9DCM/AT-Block35.2±2.7*160±13*+578±4865.7±1.5*296±6*p<0.05 vs CON+p<0.05 vs DCMWith DCM, RM (-71±l* vs -78±1mV) and APD90 (257±9* vs 226±7ms) increased, and Vmax decreased (121±5* vs 158±9V/s) compared to CON. In contrast, with AT-BLOCK, RM became more negative (-76±1+mV), APD90 was reduced (183±14*+) and Vmax increased (165±13+).SummaryAT1 AT-II receptor blockade during the progression of DCM caused significant changes in LV myocardial conduction and myocyte action potentials. These results suggest that AT1 AT-II receptor activation plays a contributory role toward the changes in LV electrophysiology with DCM

HIV Drug Resistance Early Warning Indicators in Cohorts of Individuals Starting Antiretroviral Therapy Between 2004 and 2009: World Health Organization Global Report From 50 Countries

The World Health Organization developed a set of human immunodeficiency virus drug resistance (HIVDR) early warning indicators (EWIs) to assess antiretroviral therapy clinic and program factors associated with HIVDR. EWIs are monitored by abstracting data routinely recorded in clinical records, and the results enable clinics and program managers to identify problems that should be addressed to minimize preventable emergence of HIVDR in clinic populations. As of June 2011, 50 countries monitored EWIs, covering 131 686 patients initiating antiretroviral treatment between 2004 and 2009 at 2107 clinics. HIVDR prevention is associated with patient care (appropriate prescribing and patient monitoring), patient behavior (adherence), and clinic/program management efforts to reduce treatment interruptions (follow up, retention on first-line ART, procurement and supply management of antiretroviral drugs). EWIs measure these factors and the results have been used to optimize patient and population treatment outcome

Economics of residential gas furnaces and water heaters in United States new construction market

New single-family home construction represents a significant and important market for the introduction of energy-efficient gas-fired space heating and water-heating equipment. In the new construction market, the choice of furnace and water-heater type is primarily driven by first cost considerations and the availability of power vent and condensing water heaters. Few analysis have been performed to assess the economic impacts of the different combinations of space and water-heating equipment. Thus, equipment is often installed without taking into consideration the potential economic and energy savings of installing space and water-heating equipment combinations. In this study, we use a life-cycle cost analysis that accounts for uncertainty and variability of the analysis inputs to assess the economic benefits of gas furnace and water-heater design combinations. This study accounts not only for the equipment cost but also for the cost of installing, maintaining, repairing, and operating the equipment over its lifetime. Overall, this study, which is focused on US single-family new construction households that install gas furnaces and storage water heaters, finds that installing a condensing or power-vent water heater together with condensing furnace is the most cost-effective option for the majority of these houses. Furthermore, the findings suggest that the new construction residential market could be a target market for the large-scale introduction of a combination of condensing or power-vent water heaters with condensing furnaces