Efficient Enzymatic Synthesis of Carbamates in Water Using Promiscuous Esterases/Acyltransferases.

Biocatalysis provides an attractive approach to facilitate synthetic reactions in aqueous media. Motivated by the discovery of promiscuous aminolysis activity of esterases, we exploited the esterase from Pyrobaculum calidifontis VA1 (PestE) for the synthesis of carbamates from different aliphatic, aromatic, and arylaliphatic amines and a set of carbonates such as dimethyl-, dibenzyl-, or diallyl carbonate. Thus, aniline and benzylamine derivatives, aliphatic and even secondary amines could be efficiently converted into the corresponding benzyloxycarbonyl (Cbz)- or allyloxycarbonyl (Alloc)-protected products in bulk water, with (isolated) yields of up to 99 %.

Chronic oxidative stress adaptation in head and neck cancer cells generates slow-cyclers with decreased tumour growth in vivo.

BACKGROUND: Reactive oxygen species (ROS) are implicated in cancer therapy and as drivers of microenvironmental tumour cell adaptations. Medical gas plasma is a multi-ROS generating technology that has been shown effective for palliative tumour control in head and neck cancer (HNC) patients before tumour cells adapted to the oxidative stress and growth regressed fatally. METHODS: In a bedside-to-bench approach, we sought to explore the oxidative stress adaptation in two human squamous cell carcinoma cell lines. Gas plasma was utilised as a putative therapeutic agent and chronic oxidative stress inducer. RESULTS: Cellular responses of single and multiple treated cells were compared regarding sensitivity, cellular senescence, redox state and cytokine release. Whole transcriptome analysis revealed a strong correlation of cancer cell adaption with increased interleukin 1 receptor type 2 (IL1R2) expression. Using magnetic resonance imaging, tumour growth and gas plasma treatment responses of wild-type (WT) and repeatedly exposed (RE) A431 cells were further investigated in a xenograft model in vivo. RE cells generated significantly smaller tumours with suppressed inflammatory secretion profiles and increased epidermal growth factor receptor (EGFR) activity showing significantly lower gas plasma sensitivity until day 8. CONCLUSIONS: Clinically, combination treatments together with cetuximab, an EGFR inhibitor, may overcome acquired oxidative stress resistance in HNC.

Marine Bacteroidetes enzymatically digest xylans from terrestrial plants.

Marine Bacteroidetes that degrade polysaccharides contribute to carbon cycling in the ocean. Organic matter, including glycans from terrestrial plants, might enter the oceans through rivers. Whether marine bacteria degrade structurally related glycans from diverse sources including terrestrial plants and marine algae was previously unknown. We show that the marine bacterium Flavimarina sp. Hel_I_48 encodes two polysaccharide utilization loci (PULs) which degrade xylans from terrestrial plants and marine algae. Biochemical experiments revealed activity and specificity of the encoded xylanases and associated enzymes of these PULs. Proteomics indicated that these genomic regions respond to glucuronoxylans and arabinoxylans. Substrate specificities of key enzymes suggest dedicated metabolic pathways for xylan utilization. Some of the xylanases were active on different xylans with the conserved ß-1,4-linked xylose main chain. Enzyme activity was consistent with growth curves showing Flavimarina sp. Hel_I_48 uses structurally different xylans. The observed abundance of related xylan-degrading enzyme repertoires in genomes of other marine Bacteroidetes indicates similar activities are common in the ocean. The here presented data show that certain marine bacteria are genetically and biochemically variable enough to access parts of structurally diverse xylans from terrestrial plants as well as from marine algal sources.

Insights in ChAdOx1 nCoV-19 vaccine-induced immune thrombotic thrombocytopenia.

SARS-CoV-2 vaccine ChAdOx1 nCoV-19 (AstraZeneca) causes a thromboembolic complication termed vaccine-induced immune thrombotic thrombocytopenia (VITT). Using biophysical techniques, mouse models, and analysis of VITT patient samples, we identified determinants of this vaccine-induced adverse reaction. Super-resolution microscopy visualized vaccine components forming antigenic complexes with platelet factor 4 (PF4) on platelet surfaces to which anti-PF4 antibodies obtained from VITT patients bound. PF4/vaccine complex formation was charge-driven and increased by addition of DNA. Proteomics identified substantial amounts of virus production-derived T-REx HEK293 proteins in the ethylenediaminetetraacetic acid (EDTA)-containing vaccine. Injected vaccine increased vascular leakage in mice, leading to systemic dissemination of vaccine components known to stimulate immune responses. Together, PF4/vaccine complex formation and the vaccine-stimulated proinflammatory milieu trigger a pronounced B-cell response that results in the formation of high-avidity anti-PF4 antibodies in VITT patients. The resulting high-titer anti-PF4 antibodies potently activated platelets in the presence of PF4 or DNA and polyphosphate polyanions. Anti-PF4 VITT patient antibodies also stimulated neutrophils to release neutrophil extracellular traps (NETs) in a platelet PF4-dependent manner. Biomarkers of procoagulant NETs were elevated in VITT patient serum, and NETs were visualized in abundance by immunohistochemistry in cerebral vein thrombi obtained from VITT patients. Together, vaccine-induced PF4/adenovirus aggregates and proinflammatory reactions stimulate pathologic anti-PF4 antibody production that drives thrombosis in VITT. The data support a 2-step mechanism underlying VITT that resembles the pathogenesis of (autoimmune) heparin-induced thrombocytopenia.

Connecting Algal Polysaccharide Degradation to Formaldehyde Detoxification.

Formaldehyde is a toxic metabolite that is formed in large quantities during bacterial utilization of the methoxy sugar 6-O-methyl-d-galactose, an abundant monosaccharide in the red algal polysaccharide porphyran. Marine bacteria capable of metabolizing porphyran must therefore possess suitable detoxification systems for formaldehyde. We demonstrate here that detoxification of formaldehyde in the marine Flavobacterium Zobellia galactanivorans proceeds via the ribulose monophosphate pathway. Simultaneously, we show that the genes encoding the key enzymes of this pathway are important for maintaining high formaldehyde resistance. Additionally, these genes are upregulated in the presence of porphyran, allowing us to connect porphyran degradation to the detoxification of formed formaldehyde.

Bioactive lipid screening during respiratory tract infections with bacterial and viral pathogens in mice.

INTRODUCTION: Respiratory tract infections are a worldwide health problem for humans and animals. Different cell types produce lipid mediators in response to infections, which consist of eicosanoids like hydroxyeicosatetraenoic acids (HETEs) or oxylipins like hydroxydocosahexaenoic acids (HDHAs). Both substance classes possess immunomodulatory functions. However, little is known about their role in respiratory infections. OBJECTIVES: Here, we aimed to analyze the lipid mediator imprint of different organs of C57BL/6J mice after intranasal mono-infections with Streptococcus pneumoniae (pneumococcus), Staphylococcus aureus or Influenza A virus (IAV) as wells as pneumococcal-IAV co-infection. METHODS: C57BL/6J mice were infected with different pathogens and lungs, spleen, and plasma were collected. Lipid mediators were analyzed using HPLC-MS/MS. In addition, spatial-distribution of sphingosine 1-phosphate (S1P) and ceramide 1-phosphates (C1P) in tissue samples was examined using MALDI-MS-Imaging. The presence of bacterial pathogens in the lung was confirmed via immunofluorescence staining. RESULTS: We found IAV specific changes for different HDHAs and HETEs in mouse lungs as well as enhanced levels of 20-HETE in severe S. aureus infection. Moreover, MALDI-MS-Imaging analysis showed an accumulation of C1P and a decrease of S1P during co-infection in lung and spleen. Long chain C1P was enriched in the red and not in the white pulp of the spleen. CONCLUSIONS: Lipid mediator analysis showed that host synthesis of bioactive lipids is in part specific for a certain pathogen, in particular for IAV infection. Furthermore, MS-Imaging displayed great potential to study infections and revealed changes of S1P and C1P in lungs and spleen of co-infected animals, which was not described before.

Systemic Bile Acids Affect the Severity of Acute Pancreatitis in Mice Depending on Their Hydrophobicity and the Disease Pathogenesis.

Acute pancreatitis (AP) is a major, globally increasing gastrointestinal disease and a biliary origin is the most common cause. However, the effects of bile acids (BAs), given systemically, on the pancreas and on disease severity remains elusive. In this study, we have investigated the roles of different circulating BAs in animal models for AP to elucidate their impact on disease severity and the underlying pathomechanisms. BAs were incubated on isolated acini and AP was induced through repetitive injections of caerulein or L-arginine; pancreatic duct ligation (PDL); or combined biliopancreatic duct ligation (BPDL). Disease severity was assessed using biochemical and histological parameters. Serum cholecystokinin (CCK) concentrations were determined via enzyme immunoassay. The binding of the CCK1 receptor was measured using fluorescence-labeled CCK. In isolated acini, hydrophobic BAs mitigated the damaging effects of CCK. The same BAs further enhanced pancreatitis in L-arginine- and PDL-based pancreatitis, whereas they ameliorated pancreatic damage in the caerulein and BPDL models. Mechanistically, the binding affinity of the CCK1 receptor was significantly reduced by hydrophobic BAs. The hydrophobicity of BAs and the involvement of CCK seem to be relevant in the course of AP. Systemic BAs may affect the severity of AP by interfering with the CCK1 receptor.

Carbon Source-Dependent Reprogramming of Anaerobic Metabolism in Staphylococcus aureus.

To be a successful pathogen, Staphylococcus aureus has to adapt its metabolism to the typically oxygen- and glucose-limited environment of the host. Under fermenting conditions and in the presence of glucose, S. aureus uses glycolysis to generate ATP via substrate-level phosphorylation and mainly lactic acid fermentation to maintain the redox balance by reoxidation of NADH equivalents. However, it is less clear how S. aureus proceeds under anoxic conditions and glucose limitation, likely representing the bona fide situation in the host. Using a combination of proteomic, transcriptional, and metabolomic analyses, we show that in the absence of an abundant glycolysis substrate, the available carbon source pyruvate is converted to acetyl coenzyme A (AcCoA) in a pyruvate formate-lyase (PflB)-dependent reaction to produce ATP and acetate. This process critically depends on derepression of the catabolite control protein A (CcpA), leading to upregulation of pflB transcription. Under these conditions, ethanol production is repressed to prevent wasteful consumption of AcCoA. In addition, our global and quantitative characterization of the metabolic switch prioritizing acetate over lactate fermentation when glucose is absent illustrates examples of carbon source-dependent control of colonization and pathogenicity factors.IMPORTANCE Under infection conditions, S. aureus needs to ensure survival when energy production via oxidative phosphorylation is not possible, e.g., either due to the lack of terminal electron acceptors or by the inactivation of components of the respiratory chain. Under these conditions, S. aureus can switch to mixed-acid fermentation to sustain ATP production by substrate level phosphorylation. The drop in the cellular NAD+/NADH ratio is sensed by the repressor Rex, resulting in derepression of fermentation genes. Here, we show that expression of fermentation pathways is further controlled by CcpA in response to the availability of glucose to ensure optimal resource utilization under growth-limiting conditions. We provide evidence for carbon source-dependent control of colonization and virulence factors. These findings add another level to the regulatory network controlling mixed-acid fermentation in S. aureus and provide additional evidence for the lifestyle-modulating effect of carbon sources available to S. aureus.

Virulence Factors Produced by Staphylococcus aureus Biofilms Have a Moonlighting Function Contributing to Biofilm Integrity.

Staphylococcus aureus is the causative agent of various biofilm-associated infections in humans causing major healthcare problems worldwide. This type of infection is inherently difficult to treat because of a reduced metabolic activity of biofilm-embedded cells and the protective nature of a surrounding extracellular matrix (ECM). However, little is known about S. aureus biofilm physiology and the proteinaceous composition of the ECM. Thus, we cultivated S. aureus biofilms in a flow system and comprehensively profiled intracellular and extracellular (ECM and flow-through (FT)) biofilm proteomes, as well as the extracellular metabolome compared with planktonic cultures. Our analyses revealed the expression of many pathogenicity factors within S. aureus biofilms as indicated by a high abundance of capsule biosynthesis proteins along with various secreted virulence factors, including hemolysins, leukotoxins, and lipases as a part of the ECM. The activity of ECM virulence factors was confirmed in a hemolysis assay and a Galleria mellonella pathogenicity model. In addition, we uncovered a so far unacknowledged moonlighting function of secreted virulence factors and ribosomal proteins trapped in the ECM: namely their contribution to biofilm integrity. Mechanistically, it was revealed that this stabilizing effect is mediated by the strong positive charge of alkaline virulence factors and ribosomal proteins in an acidic ECM environment, which is caused by the release of fermentation products like formate, lactate, and acetate because of oxygen limitation in biofilms. The strong positive charge of these proteins most likely mediates electrostatic interactions with anionic cell surface components, eDNA, and anionic metabolites. In consequence, this leads to strong cell aggregation and biofilm stabilization. Collectively, our study identified a new molecular mechanism during S. aureus biofilm formation and thus significantly widens the understanding of biofilm-associated S. aureus infections - an essential prerequisite for the development of novel antimicrobial therapies.

Primary and secondary gliosarcoma: differences in treatment and outcome.

INTRODUCTION: There are only few studies comparing differences in the outcome of primary versus secondary gliosarcoma. This study aimed to review the outcome and survival of patients with primary or secondary gliosarcoma following surgical resection and adjuvant treatment. The data were also matched with data of patients with primary and secondary glioblastoma (GBM). PATIENTS AND METHODS: Treatment histories of 10 patients with primary gliosarcoma and 10 patients with secondary gliosarcoma were analysed and compared. Additionally, data of 20 patients with primary and 20 patients with secondary GBM were analysed and compared. All patients underwent surgical resection of the tumour in our department. Follow-up data, progression-free survival (PFS), and median overall survival (mOS) were evaluated. RESULTS: The median PFS in patients with primary gliosarcoma was significantly higher than in patients with secondary gliosarcoma (p = 0.037). The 6-month PFS rates were 80.0% in patients with primary and 30.0% in patients with secondary gliosarcoma. Upon recurrence, five patients with primary gliosarcoma and four patients with secondary gliosarcoma underwent repeat surgical resection. The mOS of patients with primary gliosarcoma was significantly higher than that of patients with secondary gliosarcoma (p = 0.031). The percentage of patients surviving at 1-year/2-year follow-up in primary gliosarcoma was 70%/20%, while it was only 10%/10% in secondary gliosarcoma. When PFS and mOS of primary gliosarcoma was compared to primary GBM, there were no statistically differences (p = 0.509; p = 0.435). The PFS and mOS of secondary gliosarcoma and secondary GBM were also comparable (p = 0.290 and p = 0.390). CONCLUSION: Patients with primary gliosarcoma have a higher PFS and mOS compared to those with secondary gliosarcoma. In the case of tumour recurrence, patients with secondary gliosarcoma harbour an unfavourable prognosis with limited further options. The outcome of patients with primary or secondary gliosarcoma is comparable to that of patients with primary or secondary GBM.

Novel Small-Molecule Hybrid-Antibacterial Agents against S. aureus and MRSA Strains.

Ongoing resistance developments against antibiotics that also affect last-resort antibiotics require novel antibacterial compounds. Strategies to discover such novel structures have been dimerization or hybridization of known antibacterial agents. We found novel antibacterial agents by dimerization of indols and hybridization with carbazoles. They were obtained in a simple one-pot reaction as bisindole tetrahydrocarbazoles. Further oxidation led to bisindole carbazoles with varied substitutions of both the indole and the carbazole scaffold. Both the tetrahydrocarbazoles and the carbazoles have been evaluated in various S. aureus strains, including MRSA strains. Those 5-cyano substituted derivatives showed best activities as determined by MIC values. The tetrahydrocarbazoles partly exceed the activity of the carbazole compounds and thus the activity of the used standard antibiotics. Thus, promising lead compounds could be identified for further studies.

Analyses of competent and non-competent subpopulations of Bacillus subtilis reveal yhfW, yhxC and ncRNAs as novel players in competence.

Upon competence-inducing nutrient-limited conditions, only part of the Bacillus subtilis population becomes competent. Here, we separated the two subpopulations by fluorescence-assisted cell sorting (FACS). Using RNA-seq, we confirmed the previously described ComK regulon. We also found for the first time significantly downregulated genes in the competent subpopulation. The downregulated genes are not under direct control by ComK but have higher levels of corresponding antisense RNAs in the competent subpopulation. During competence, cell division and replication are halted. By investigating the proteome during competence, we found higher levels of the regulators of cell division, MinD and Noc. The exonucleases SbcC and SbcD were also primarily regulated at the post-transcriptional level. In the competent subpopulation, yhfW was newly identified as being highly upregulated. Its absence reduces the expression of comG, and has a modest, but statistically significant effect on the expression of comK. Although expression of yhfW is higher in the competent subpopulation, no ComK-binding site is present in its promoter region. Mutants of yhfW have a small but significant defect in transformation. Metabolomic analyses revealed significant reductions in tricarboxylic acid (TCA) cycle metabolites and several amino acids in a ΔyhfW mutant. RNA-seq analysis of ΔyhfW revealed higher expression of the NAD synthesis genes nadA, nadB and nadC.

Escherichia coli mediated resistance of Entamoeba histolytica to oxidative stress is triggered by oxaloacetate.

Amebiasis, a global intestinal parasitic disease, is due to Entamoeba histolytica. This parasite, which feeds on bacteria in the large intestine of its human host, can trigger a strong inflammatory response upon invasion of the colonic mucosa. Whereas information about the mechanisms which are used by the parasite to cope with oxidative and nitrosative stresses during infection is available, knowledge about the contribution of bacteria to these mechanisms is lacking. In a recent study, we demonstrated that enteropathogenic Escherichia coli O55 protects E. histolytica against oxidative stress. Resin-assisted capture (RAC) of oxidized (OX) proteins coupled to mass spectrometry (OX-RAC) was used to investigate the oxidation status of cysteine residues in proteins present in E. histolytica trophozoites incubated with live or heat-killed E. coli O55 and then exposed to H2O2-mediated oxidative stress. We found that the redox proteome of E. histolytica exposed to heat-killed E. coli O55 is enriched with proteins involved in redox homeostasis, lipid metabolism, small molecule metabolism, carbohydrate derivative metabolism, and organonitrogen compound biosynthesis. In contrast, we found that proteins associated with redox homeostasis were the only OX-proteins that were enriched in E. histolytica trophozoites which were incubated with live E. coli O55. These data indicate that E. coli has a profound impact on the redox proteome of E. histolytica. Unexpectedly, some E. coli proteins were also co-identified with E. histolytica proteins by OX-RAC. We demonstrated that one of these proteins, E. coli malate dehydrogenase (EcMDH) and its product, oxaloacetate, are key elements of E. coli-mediated resistance of E. histolytica to oxidative stress and that oxaloacetate helps the parasite survive in the large intestine. We also provide evidence that the protective effect of oxaloacetate against oxidative stress extends to Caenorhabditis elegans.

Biotransformation of bisphenol A analogues by the biphenyl-degrading bacterium Cupriavidusbasilensis - a structure-biotransformation relationship.

Comparative analyses determined the relationship between the structure of bisphenol A (BPA) as well as of seven bisphenol analogues (bisphenol B (BPB), bisphenol C (BPC), bisphenol E (BPE), bisphenol F (BPF), bisphenol Z (BPZ), bisphenol AP (BPAP), bisphenol PH (BPPH)) and their biotransformability by the biphenyl-degrading bacterium Cupriavidus basilensis SBUG 290. All bisphenols were substrates for bacterial transformation with conversion rates ranging from 6 to 98% within 216 h and 36 different metabolites were characterized. Transformation by biphenyl-grown cells comprised four different pathways: (a) formation of ortho-hydroxylated bisphenols, hydroxylating either one or both phenols of the compounds; (b) ring fission; (c) transamination followed by acetylation or dimerization; and (d) oxidation of ring substituents, such as methyl groups and aromatic ring systems, present on the 3-position. However, the microbial attack of bisphenols by C. basilensis was limited to the phenol rings and its substituents, while substituents on the carbon bridge connecting the rings were not oxidized. All bisphenol analogues with modifications at the carbon bridge could be oxidized up to ring cleavage, while substituents at the 3-position of the phenol ring other than hydroxyl groups did not allow this reaction. Replacing one methyl group at the carbon bridge of BPA by a hydrophobic aromatic or alicyclic ring system inhibited both dimerization and transamination followed by acetylation. While most of the bisphenol analogues exhibited estrogenic activity, four biotransformation products tested were not estrogenically active.

Metabolic inventory of Streptococcus pneumoniae growing in a chemical defined environment.

The Gram-positive bacterium Streptococcus pneumoniae can cause a broad range of severe diseases including pneumonia and septicemia. The pneumococcal pathophysiology is highly dependent on host nutrients such as purines, pyrimidines, amino acids and carbon sources. Therefore, we aimed to decipher the metabolome with a metabolomics approach that allows for the investigation of the basic metabolic characteristics during growth in a chemical defined medium composed of typical host metabolites. By using a combination of 1H-NMR, HPLC-MS and GC-MS methods we monitored extracellular uptake and secretion of metabolites as well as the intracellular metabolic composition. Employing our validated protocol for the pneumococcal intracellular metabolome analysis, a time resolved snapshot of the primary metabolism of pneumococci was obtained. The intracellular metabolic profile indicates a high glycolytic flux and displays high concentrated precursors of peptidoglycan synthesis probably to fuel cell-wall-metabolism in growing cells. Furthermore, our data reflect the biochemical dependency for S. pneumoniae on external host derived nutrients such as nucleosides. These essential pathways may serve as new targets in the drug development against S. pneumoniae.

Cerebellar glioblastoma: a clinical series with contemporary molecular analysis.

BACKGROUND: Glioblastomas (GBM) are localized in only less than 1% of patients in the cerebellum. Therefore, tumor characteristics, survival, and the efficacy of therapies are not yet well defined. The present study aims to characterize the molecular features of cerebellar GBM (GBMc) in 8 patients treated with contemporary modality in our institution. METHODS: Patients' treatment history, progression-free survival (PFS), and overall survival (OS) were analyzed. All histopathological specimens were re-investigated. EGFR amplification was determined by FISH, H3F3A, and HIST1H3B mutation status and MGMT promoter methylation after bisulfite treatment by pyrosequencing and BRAF V600E by pyrosequencing and immunohistochemistry. TERT promoter mutations were analyzed by Sanger sequencing, CDKN2A/B deletions by digital PCR. The expression of IDH1 R132H, ATRX, and p53 was determined through immunohistochemistry. RESULTS: Six adults and two children (mean age 36 years) underwent tumor resection via medial or lateral suboccipital craniotomy. The median overall survival (mOS) of the adult patients was 7 months. GBMc from two children demonstrated a H3F3A K27M mutation. One of these also harbored a BRAF V600E mutation and has already had a PFS of 74 months. Mutated IDH1 R132H protein was expressed in 2 GBM from adult patients with previous supratentorial anaplastic astrocytoma. One patient carried a TERT promoter mutation. Another patient initially presented with a thalamic pilocytic astrocytoma. The cerebellar tumor revealed neither a BRAF V600E nor a H3F3A mutation but a homozygous CDKN2A/B deletion. CONCLUSIONS: GBM located in the cerebellum can be found in all age groups. We provide novel molecular genetic data on these rare tumors. Mutated IDH1 R132H protein and H3F3A K27M mutations indicate that a substantial number of GBMc are "metastatic" or "diaschismatic" lesions. Mutation of BRAF V600E may have a stronger biological significance than H3F3A K27M alterations. In a subset of patients, GBM may arise primarily as a distinct entity in the cerebellum.

Influence of soy isoflavones in breast cancer angiogenesis: a multiplex glass ELISA approach.

PURPOSE: The aim of this study was to evaluate the anti-angiogenic properties of soy isoflavones using two breast cancer cell lines, by measuring the concentration of 30 cytokines involved in angiogenesis using a multiplex glass slide ELISA-based array. METHODS: Estrogen-dependent MCF-7 cells and estrogen-independent MDA-MB-231 cells were exposed to genistein (Gen), daidzein (Dai) and a soy seed extract (Ext) for 72 hrs, at selected concentration levels. The conditioned medium was analyzed using a glass slide, multiplex sandwich ELISA-based platform with fluorescent detection which allowed the identification and the quantification of 30 angiogenesis-related cytokines. RESULTS: In MCF-7 cells, low, stimulatory concentrations of test compounds determined the increase of CXCL16 and VEGF-A level. Gen induced the greatest effect, with 1.5-fold change compared to control. When MDA-MB-231 cells were exposed to inhibitory concentrations, all test compounds determined a reduction of CXCL16 and VEGF-A level with approximately 30%. CONCLUSIONS: Soluble CXCL16 and VEGF-A are two promoters of angiogenesis and metastasis in breast cancer. The stimulation of these two angiogenesis-related cytokines could represent one of the mechanisms explaining the proliferative effects of low isoflavone doses in estrogen-dependent cells. In estrogen-independent cells, soy isoflavones inhibited their secretion, demonstrating promising anti-angiogenic properties.

Biotransformation and reduction of estrogenicity of bisphenol A by the biphenyl-degrading Cupriavidus basilensis.

The biphenyl-degrading Gram-negative bacterium Cupriavidus basilensis (formerly Ralstonia sp.) SBUG 290 uses various aromatic compounds as carbon and energy sources and has a high capacity to transform bisphenol A (BPA), which is a hormonally active substance structurally related to biphenyl. Biphenyl-grown cells initially hydroxylated BPA and converted it to four additional products by using three different transformation pathways: (a) formation of multiple hydroxylated BPA, (b) ring fission, and (c) transamination followed by acetylation or dimerization. Products of the ring fission pathway were non-toxic and all five products exhibited a significantly reduced estrogenic activity compared to BPA. Cell cultivation with phenol and especially in nutrient broth (NB) resulted in a reduced biotransformation rate and lower product quantities, and NB-grown cells did not produce all five products in detectable amounts. Thus, the question arose whether enzymes of the biphenyl degradation pathway are involved in the transformation of BPA and was addressed by proteomic analyses.

Novel Effective Small-Molecule Antibacterials against Enterococcus Strains.

Enterococcus species cause increasing numbers of infections in hospitals. They contribute to the increasing mortality rates, mostly in patients with comorbidities, who suffer from severe diseases. Enterococcus resistances against most antibiotics have been described, including novel antibiotics. Therefore, there is an ongoing demand for novel types of antibiotics that may overcome bacterial resistances. We discovered a novel class of antibiotics resulting from a simple one-pot reaction of indole and o-phthaldialdehyde. Differently substituted indolyl benzocarbazoles were yielded. Both the indole substitution and the positioning at the molecular scaffold influence the antibacterial activity towards the various strains of Enterococcus species with the highest relevance to nosocomial infections. Structure-activity relationships are discussed, and the first lead compounds were identified as also being effective in the case of a vancomycin resistance.

A proteomic approach for the identification of immunotoxic properties of Tulipalin A.

The immune system is permanently exposed to several environmental influences that can have adverse effects on immune cells or organs leading to immunosuppression or inappropriate immunostimulation, called direct immunotoxicity. The natural compound Tulipalin A (TUPA), a lactone with α-methylene-γ-butyrolactone moiety, can influence the immune system and lead to allergic contact dermatitis. This in vitro study focused on effects of TUPA using two immune cell lines (Jurkat T cells and THP-1 monocytes). To evaluate the immunotoxic potential of the compound, a proteomic approach applying 2D gel electrophoresis and MALDI-TOF/TOF-MS in combination with metabolomic analysis was used after exposure of the cells to IC10 of TUPA. THP-1 cells showed a strong robustness to TUPA treatment since only five proteins were altered. In contrast, in Jurkat T cells an increase in the abundance of 66 proteins and a decrease of six proteins was determined. These intracellular proteins were mapped to biological processes. Especially an accumulation of chaperones and an influence on the purine synthesis were observed. The changes in purine synthesis were confirmed by metabolomic analysis. In conclusion, the data indicate possible target processes of low doses of TUPA in Jurkat T cells and provides knowledge of how TUPA affects the functionality of immune cells.