ABSTRACT: Prevotella bryantii B14 was cultivated with monensin. Growth was monitored over a period of 9h with a broad range of monensin concentrations.
Project description:Prevotella bryantii B14 was cultivated with monensin. Growth was monitored over a period of 72h including frequent sampling of cells.
Project description:P. bryantii B14 cells were cultivated separately in acetic (Acet), propionic (Prop), butyric (But), iso-butyric (iBut), valeric (Val), iso-valeric (iVal) and 2-methyl butyric acid (2MB) as well as in a mixture of all mentioned short-chain fatty acids (Mix). All 8 treatments were analyzed regarding their proteomes in order to understand the requirements and effects of each SCFA on the metabolism.
Project description:The aim of the present work was to investigate the effect of monensin on the in vitro growth of T. gondii tachyzoites and on the host cells (human brain microvascular endothelial cells - hBMECs). The hypotheses were that (1) inhibition of the WNT signalling pathway by monensin can reduce the growth of T. gondii infecting human brain microvascular endothelial cells (hBMECs) and (2) by suppression of the growth of T. gondii using monensin, impairment of the BBB integrity can be restored (3) inhibition of WNT pathway by monensin can be detected by microarray experiment.
Project description:Blood samples from healthy controls and PAD patients were collected at Xiangya Hospital, China. Two to three milliliters of venous blood were drawn into EDTA-coated tubes and stored at 4°C. Plasma was isolated after centrifugation (2,000 g, 10 min, 4°C), aliquoted, and send to perform MS. Group1: Health control D1, A5, A46, B14,B22; Group2: PAD C6 C29 D45 A32
Project description:Prevotella intermedia is a Gram-negative bacterium that is notably linked to periodontitis and acute necrotizing ulcerative gingivitis. P. intermedia is known to utilise the T9SS to secrete and anchor virulence factors to the cell surface, presumably via C-terminal modification of the cargo protein with a cell surface polysaccharide. The identity of the linking sugar and the sites of modification on the cargo have yet to be determined. Here, we first employed Hidden Markov Models to predict cargo proteins in P. intermedia, and then conducted LC-MS/MS analyses of partially deglycosylated fractions to characterise the C-terminal glycosylation. A total of 80 cargo proteins were predicted based on the presence of a T9SS C-terminal domain signal (CTD), and these were divided into 48 short CTDs predicted by AlphaFold to adopt a single 3-stranded β-sheet and 32 long CTDs predicted to form the more typical larger β-sandwich structures. Cleavage sites for five short CTDs and four long CTDs were experimentally determined, and glycosylation was observed at the mature C-terminus of six cargo. Two glycans were identified of delta masses 419.198 and 433.185 Da, corresponding to novel linkages to N-alanyl dHex-HexNAc and N-alanyl (Me-dHex)-HexNAc, respectively, with the alanine amide-linked to the protein C-terminus. This indicated that both short and long CTDs were cleaved and glycosylated. AlphaFold multimer modelling predicted that both kinds of CTDs could bind to the PorV shuttle protein in the same manner, with the conserved CTD motifs interacting with the same sites in PorV.
Project description:A549 cells were treated for 8 or 20 h using 20 ng/mL brefeldin A (BFA), 5 µM golgicide A (GCA), 10 µM monensin (MON) or the appropriate vehicle control (BFA/EtOH, MON/EtOH, GCA/DMSO).
Project description:During radiotherapy, X-rays can deliver significant doses of ionising radiation to both cancerous and surrounding healthy tissues, often leading to undesirable side effects that compromise patient outcomes. While the cellular effects of such therapeutic x-ray exposures are well studied, the impact on extracellular matrix (ECM) proteins, which are critical to tissue integrity, remains poorly understood. This study characterises the response of breast proteins, including the major ECM components collagen I and fibronectin, to therapeutic X-rays doses of 50Gy (as employed in breast radiotherapy) and 100Gy, using a combination of gel electrophoresis, biochemical assays, and mass spectrometry-based peptide location fingerprinting (PLF) analysis. In purified protein solutions, X-ray exposure led to fragmentation of constituent α chains of collagen I, and in fibronectin localised structural modifications (as detected by LC-MS/MS peptide location fingerprinting [PLF]) which increased its binding affinity for collagen I. In complex environments, such as newly synthesised fibroblast-derived ECM (fECM) and mature ex vivo breast tissue, therapeutic x-rays induced proteolytic changes in, not just collagen I and fibronectin, but also key basement membrane proteins, including collagen IV, laminin, nidogen-1, and perlecan. Intracellular proteins associated with gene expression (RPS3, MeCP2), cytoskeleton (moesin, plectin), and the endoplasmic reticulum (calnexin) were also found to be structurally compromised. These structural changes may impair the ECM integrity and alter cell-ECM interactions, with potential implications for tissue stiffening, fibrosis, and impaired wound healing in irradiated tissues.