Project description:Native metabolomics method validation for chymotrypsin.
1. Limit of detection Mollasamide- Chymotrypsin
2. Flowinjection of Mollasamide over UHPLC gradients (with make-up).
3. Binding tests with chymotrypsin and different standards.
Project description:1) Analysis of HTPS FT (analysis of the background, without protease) 2) Analysis of specificity and activity of Thrombin and Chymotrypsin at multiple time point (0,5,15,30,60,120,240 min) 3) Analysis of specificity and activity of Thrombin and Chymotrpysin with 2 hr incubation at 20C, 25C and 37C
Project description:To investigate the global molecular mechanisms underlying the phenotype induced by ZZEF1 loss in pancreatic β-cells and subsequent treatment with azoramide, we isolated pancreatic islets from β-cell-specific ZZEF1 knockout mice (βZKO-Mip) and their littermate controls. Both groups were fed a HFD for four weeks and treated with either azoramide (10 µM) or DMSO (10 µM) for 24 hours.
Project description:Native metabolomics method validation for chymotrypsin.
1. Limit of detection Mollasamide- Chymotrypsin
2. Flowinjection of Mollasamide over UHPLC gradients (with make-up).
3. Binding tests with chymotrypsin and different standards.
Project description:Members of the serpin (serine protease inhibitor) superfamily have been identified in higher, multicellular eukaryotes, as well as in bacteria, although surveillance of available genome sequences indicates that bacterial serpin-encoding (ser) homologs are not widely distributed. In members of the genus Bifidobacterium this gene appears to be present in at least five, and perhaps up to nine, out of 30 species tested. Moreover, phylogenetic analysis using available bacterial and eukaryotic serpin sequences revealed that bifidobacteria specify serpins that form a separate clade. We characterized the ser210B locus of Bifidobacterium breve 210B, which consists of a number of genes, whose deduced protein products display significant similarity to proteins encoded by corresponding loci found in several other bifidobacteria. Northern hybridization, primer extension, micro array analysis, RT-PCR and Quantitative Real Time (qRT) - PCR analysis revealed that a 3.5 kb polycistronic mRNA, encompassing the ser210B operon with a single transcriptional start site, is strongly induced following treatment of B. breve 210B cultures with particular proteases. In contrast, transcription of the ser homolog of other bifidobacteria, such as Bifidobacterium longum subsp. infantis, Bifidobacterium dentium and B. longum subsp. longum, appears to be triggered by a different set of proteases Transcriptional response to protease treatments (kallikrein, papain and chymotrypsin) of Bifidobacterium breve 210B
Project description:Myeloid cell lines (K562 and HEL) were treated overnight with CHIR99021 or a vehicle control (DMSO). CHIR99021 treatment inhibits GSK3B within the destruction complex functioning in Wnt/beta-catenin signalling pathway, thereby preventing beta-catenin degradation and promoting its stabilization. Following the overnight incubation, beta-catenin RIP (RNA immunoprecipitation) was performed in both K562 and HEL cells. RNA samples obtained from beta-catenin RIP in these cells were then sequenced to identify beta-catenin-associated RNAs under CHIR99021 treatment compared to basal conditions (DMSO control).
Project description:mESCs were cultured at least 2 weeks in medium medium: N2B27 for SILAC (DMEM/F12 for SILAC (AthenaES), Neurobasal for SILAC (AthenaES), Sodium Pyruvate (40 mg/mL), N2 (1X), B27 (0.5X), Pen/Strep (1%), L-glutamine (2 mM), beta-mercaptoethanol (50 µM)) + 2i/LIF supplemented with (medium) 13C6 15N4 L-arginine (0.65 mM), (medium) 13C6 L-arginine (0.55 mM), and L-Proline (200 mg/L)84. Before switching from light to heavy medium, cells were treated with 0.05 µg/mL CHX or equivalent dilution of DMSO in 6-well plates. We treated the cells with 0.05 µg/mL CHX. At timepoint 0 h, we replaced medium medium with pre-warmed heavy medium: N2B27 for SILAC (Thermo scientific) supplemented with (heavy) 13C6 15N2 L-Lysine-2HCl (0.65 mM), (heavy) 13C6 15N4 L-Arginine-HCl (0.55 mM), and L-Proline (200 mg/L) for mESC.
Project description:Effects of pioglitazone and beta-naphtoflavone treatmend for 72 hours on human primary trophoblasts were investigated. 0.25% DMSO was a control.