Estimation of the genetic potential of the production of secondary metabolites from the myxobacterium Sorangium cellulosum So ce90 and identification of the spirangiene biosynthetic gene cluster.

Myxobakterien sind gram-negative, ubiquitär vorkommende Bodenbakterien. Inzwischen wurden sie als reiche Quelle für bioaktive Naturstoffe beschrieben. Aus dem Myxobakterium Sorangium cellulosum So ce90 konnten bisher zwei aktive Naturstoffe isoliert werden. Die Epothilone und die Spirangiene. Diese werden von Multienzymkomplexen biosynthetisiert, den Polyketidsynthasen (PKS) und den nicht-ribosomalen Peptidsynthetasen (NRPS). Im Verlauf dieser Arbeit konnten acht unabhängige Genloci identifiziert werden, die für PKS und/oder NRPS kodieren, darunter das Epothilon- und das Spirangienbiosynthesegencluster. Die Funktion der restlichen Genloci ist unbekannt. Durch Geninaktivierungsexperimente, die mit dem mobilisierbaren Vektor pSUPHyg und Hygromycin B als Selektionsmarker durchgeführt wurden, konnte eine sequenzierte Genregion von 48 kb als Teil des Spirangienbiosynthsegenclusters identifiziert werden. Das Gen spiL, welches eine Cytochrom P450 abhängige Monooxygenase kodiert, konnte ebenfalls in seiner Funktion charakterisiert werden. An der Regulation der Biosynthese beteiligt ist das Genprodukt von spiZ, das Ähnlichkeit zu Sigma-Faktoren der ECF-Familie zeigt. Sequenzwiederhohlungen innerhalb des Spirangienbiosynthesegenclusters lassen auf eine Evolution durch Duplizierung modulkodierender DNA-Bereiche schließen.Myxobacteria are gram-negative ubiquitous soil bacteria. There are also prominent as a rich source of bioactive natural compounds. The myxobacterium Sorangium cellulosum So ce90 produces Epothilones and the Spirangienes. They are biosynthesised by multi-enzyme complexes, called polyketidesynthases (PKS) and non-ribosomal polypeptidesynthetases (NRPS). During this research eight independent gene loci coding for PKS and/or NRPS where identified, including the biosynthetic gene clusters for the production of Epothilone and Spirangiene. The function of the other six gene loci is yet unknown. Gene inactivation experiments, using a mobiliziable vector pSUPHyg and the selection marker Hygromycin B where used to identify a part of the genes of the spirangiensynthase, a sequenzed region of about 48 kb. A gene, named spiL, which encodes a cytochrome P450 monooxygenase, was also characterized by inactivaiton, and further spiZ, that, as protein, shows high similarity to the ecf-sigma-factors. Long DNA parts within the genes coding for spirangiensynthase, where shown to be direct repeats and gave rise to the consumption of the evolutionary progress in the development of the genes by duplication processes

Human Monoclonal Antibody HCV1 Effectively Prevents and Treats HCV Infection in Chimpanzees

Hepatitis C virus (HCV) infection is a leading cause of liver transplantation and there is an urgent need to develop therapies to reduce rates of HCV infection of transplanted livers. Approved therapeutics for HCV are poorly tolerated and are of limited efficacy in this patient population. Human monoclonal antibody HCV1 recognizes a highly-conserved linear epitope of the HCV E2 envelope glycoprotein (amino acids 412-423) and neutralizes a broad range of HCV genotypes. In a chimpanzee model, a single dose of 250 mg/kg HCV1 delivered 30 minutes prior to infusion with genotype 1a H77 HCV provided complete protection from HCV infection, whereas a dose of 50 mg/kg HCV1 did not protect. In addition, an acutely-infected chimpanzee given 250 mg/kg HCV1 42 days following exposure to virus had a rapid reduction in viral load to below the limit of detection before rebounding 14 days later. The emergent virus displayed an E2 mutation (N415K/D) conferring resistance to HCV1 neutralization. Finally, three chronically HCV-infected chimpanzees were treated with a single dose of 40 mg/kg HCV1 and viral load was reduced to below the limit of detection for 21 days in one chimpanzee with rebounding virus displaying a resistance mutation (N417S). The other two chimpanzees had 0.5-1.0 log(10) reductions in viral load without evidence of viral resistance to HCV1. In vitro testing using HCV pseudovirus (HCVpp) demonstrated that the sera from the poorly-responding chimpanzees inhibited the ability of HCV1 to neutralize HCVpp. Measurement of antibody responses in the chronically-infected chimpanzees implicated endogenous antibody to E2 and interference with HCV1 neutralization although other factors may also be responsible. These data suggest that human monoclonal antibody HCV1 may be an effective therapeutic for the prevention of graft infection in HCV-infected patients undergoing liver transplantation

Quantifying Product Line Benefits

Software product lines promise benefits like development and maintenance effort reduction, time to market decrease, and quality improvement, all resulting from planned and systematic reuse of common core assets. However, very little quantitative data has been measured so far to prove these promises

Ru/Ni Dual Catalytic Desulfinative Photoredox C_sp²–C_sp³ Cross‑Coupling of Alkyl Sulfinate Salts and Aryl Halides

A mild Ru/Ni dual catalytic desulfinative photoredox C_sp²–C_sp³ cross-coupling reaction of alkyl sulfinate salts with aryl halides has been developed. The optimized catalyst system, consisting of Ru­(bpy)₃Cl₂, Ni­(COD)₂, and DBU, smoothly mediates the coupling of a diverse set of secondary and primary nonactivated alkyl sulfinate salts with a broad range of electron-deficient aryl bromides, electron-rich aryl iodides, and heteroaryl bromides under irradiation with blue light. The procedure is ideal for late-stage introduction of alkyl groups on pharmaceutical intermediates, and the C_sp²–C_sp³ cross-coupling reaction allowed the rapid synthesis of caseine kinase 1δ inhibitor analogues via a parallel medicinal chemistry effort

Serum anti-toxin B antibody correlates with protection from recurrent Clostridium difficile infection (CDI)

BACKGROUND: Previous studies have demonstrated a correlation between Clostridium difficile anti-toxin A serum antibodies and protection against symptomatic disease and recurrence. METHODS: A neutralizing monoclonal antibody to C. difficile toxin A (CDA1) developed by MBL and Medarex, Inc. was studied in a phase II, randomized, double-blind, placebo-controlled trial in patients receiving standard of care treatment for C. difficile infection (CDI). Twenty-nine subjects received a single intravenous infusion of 10mg/kg CDA1 and 17 subjects received placebo and were evaluated for recurrence of CDI during the 56-day study period. Serum antibodies against C. difficile toxin A and B were measured by ELISA and cytotoxicity assay at various time points before and after infusion. FINDINGS: CDI recurrence occurred in 5 of 29 (17%) in the CDA1 group and 3 of 17 (18%) (p=NS) in the placebo group with a trend toward delay in time to recurrence in the group treated with CDA1. The geometric mean concentration of antibody to an epitope of the receptor-binding domain of toxin B (0.300 and 1.20microg/ml, respectively; p=0.02) and geometric mean titer of neutralizing B antibody (8.00 and 100, respectively; p=0.02) at study day 28 were lower for those subjects with recurrence compared to those who did not recur. In addition, a significantly greater proportion of subjects who recurred were infected with the epidemic BI/NAP1/027 strain compared with those that did not recur (88% vs. 22%; p=0.002). Finally, in a multiple logistic regression analysis neutralizing anti-toxin B at day 14 (p\u3c0.001), anti-toxin A at day 28 (p\u3c0.001) and infection with the BI/NAP1/027 strain at enrollment (p=0.002) were all predictive of CDI recurrence. INTERPRETATION: In this prospective study, lower concentrations of neutralizing anti-toxin B and anti-toxin A antibody and infection with the BI/NAP1/027 strain of C. difficile were significantly associated with recurrence of CDI

Ligand-to-Copper Charge Transfer: A General Catalytic Approach to Aromatic Decarboxylative Functionalization

Aryl carboxylic acids are valuable, stable, and abundant functional handles in organic synthesis. Historically, their activation with established two-electron methods requires forcing conditions, and such protocols are limited in scope. In contrast, we envisioned that copper’s ability to generate open-shell species through ligand-to-metal charge transfer (LMCT), combined with its unique capacity to act as a potential aroyloxy and aryl radical reservoir, could mediate facile light- and copper-enabled aromatic decarboxylative functionalization by mitigating undesired reactivity of radical intermediates formed during aromatic decarboxylation. We report herein a general copper-LMCT open-shell activation platform for aromatic halodecarboxylation. Catalytic decarboxylative chlorination, bromination, and iodination of diverse (hetero)aryl carboxylic acids have been achieved to provide broadly used electrophilic cross-coupling handles from widely available aromatic acid precursors. Notably, decarboxylative fluorination of aryl carboxylic acids ­– a long-standing challenge in the field of organic synthesis – is readily accessible over a wide breadth of (hetero)aryl substrates. Ultrafast transient absorption (TA) spectroscopy experiments in combination with steady-state UV-vis spectroscopy studies are consistent with the proposed copper-LMCT mechanism, supporting the mechanistic basis of this activation platform.</p

Copper-Mediated Perfluoroalkylation of Heteroaryl Bromides with (phen)CuR F

[Image: see text] The attachment of perfluoroalkyl groups onto organic compounds has been a major synthetic goal over the past several decades. Previously, our group reported phenanthroline-ligated perfluoroalkyl copper reagents, (phen)CuR(F), which react with aryl iodides and aryl boronates to form the corresponding benzotrifluorides. Herein the perfluoroalkylation of a series of heteroaryl bromides with (phen)CuCF(3) and (phen)CuCF(2)CF(3) is reported. The mild reaction conditions allow the process to tolerate many common functional groups. Perfluoroethylation with (phen)CuCF(2)CF(3) occurs in somewhat higher yields than trifluoromethylation with (phen)CuCF(3), creating a method to generate fluoroalkyl heteroarenes that are less accessible from trifluoroacetic acid derivatives