Search results for "Porters"
showing 10 items of 233 documents
Real-Time RT-PCR assay to quantify the expression of fum1 and fum19 genes from the Fumonisin-producing Fusarium verticillioides
2007
Fumonisins are a group of mycotoxins produced by Fusarium species of the Gibberella fujikuroi species complex that contaminate food and feed products, and represent a risk for human and animal health. In this work, we have developed a specific real-time reverse transcription-PCR (RT-PCR) assay to quantify the level of expression of two genes of the fumonisin biosynthetic cluster in F. verticillioides: fum1 (that encodes a polyketide synthase enzyme) and the ABC transporter encoding gene fum19. The level of expression of both genes was compared with the amount of fumonisin B(1) (FB(1)), measured by HPLC, produced by several strains of F. verticillioides in liquid culture. The results indicat…
Transcriptional expression of selected genes associated with excretion of carboxylic acids from aci mutants of Saccharomyces cerevisiae
2013
Introduction: Saccharomyces cerevisiae is an excellent model organism for studies of transcriptional regulation of metabolic processes in other eukaryotic cells including human cells. Cellular acid-base balance can be disturbed in pathologic situations such as renal acidosis or cancer. The extracellular pH of malignant solid tumors is acidic in the range of 6.5-6.9. EG07 and EG37 aci mutants of Saccharomyces cerevisiae excessively excrete carboxylic acids to glucose-containing media or distilled water. The excreted acids are Krebs and/or glyoxylate cycle intermediates. The genes restoring the wild-type phenotype have function that does not easily explain theAci phenotype.Material/Methods: I…
Functional cysteine-less subunits of the transporter associated with antigen processing (TAP1 and TAP2) by de novo gene assembly
2002
AbstractWithin the adaptive immune system the transporter associated with antigen processing (TAP) plays a pivotal role in loading of peptides onto major histocompatibility (MHC) class I molecules. As a central tool to investigate the structure and function of the TAP complex, we created cysteine-less human TAP subunits by de novo gene synthesis, replacing all 19 cysteines in TAP1 and TAP2. After expression in TAP-deficient human fibroblasts, cysteine-less TAP1 and TAP2 are functional with respect to adenosine triphosphate (ATP)-dependent peptide transport and inhibition by ICP47 from herpes simplex virus. Cysteine-less TAP1 and TAP2 restore maturation and intracellular trafficking of MHC c…
Distant downstream sequence determinants can control N-tail translocation during protein insertion into the endoplasmic reticulum membrane.
2000
We have studied the membrane insertion of ProW, an Escherichia coli inner membrane protein with seven transmembrane segments and a large periplasmic N-terminal tail, into endoplasmic reticulum (ER)-derived dog pancreas microsomes. Strikingly, significant levels of N-tail translocation is seen only when a minimum of four of the transmembrane segments are present; for constructs with fewer transmembrane segments, the N-tail remains mostly nontranslocated and the majority of the molecules adopt an 'inverted' topology where normally nontranslocated parts are translocated and vice versa. N-tail translocation can also be promoted by shortening of the N-tail and by the addition of positively charg…
The Nature of the Stimulus and of the Fumarate Binding Site of the Fumarate Sensor DcuS of Escherichia coli
2005
DcuS is a membrane-associated sensory histidine kinase of Escherichia coli specific for C(4) -dicarboxylates. The nature of the stimulus and its structural prerequisites were determined by measuring the induction of DcuS-dependent dcuB'-'lacZ gene expression. C(4)-dicarboxylates without or with substitutions at C2/C3 by hydrophilic (hydroxy, amino, or thiolate) groups stimulated gene expression in a similar way. When one carboxylate was replaced by sulfonate, methoxy, or nitro groups, only the latter (3-nitropropionate) was active. Thus, the ligand of DcuS has to carry two carboxylate or carboxylate/nitro groups 3.1-3.8 A apart from each other. The effector concentrations for half-maximal i…
Antigen processing influences HIV-specific cytotoxic T lymphocyte immunodominance
2016
Udgivelsesdato: 2009-May-03 Although cytotoxic T lymphocytes (CTLs) in people infected with human immunodeficiency virus type 1 can potentially target multiple virus epitopes, the same few are recognized repeatedly. We show here that CTL immunodominance in regions of the human immunodeficiency virus type 1 group-associated antigen proteins p17 and p24 correlated with epitope abundance, which was strongly influenced by proteasomal digestion profiles, affinity for the transporter protein TAP, and trimming mediated by the endoplasmatic reticulum aminopeptidase ERAAP, and was moderately influenced by HLA affinity. Structural and functional analyses demonstrated that proteasomal cleavage 'prefer…
Topology and accessibility of the transmembrane helices and the sensory site in the bifunctional transporter DcuB of Escherichia coli.
2011
C(4)-Dicarboxylate uptake transporter B (DcuB) of Escherichia coli is a bifunctional transporter that catalyzes fumarate/succinate antiport and serves as a cosensor of the sensor kinase DcuS. Sites and domains of DcuB were analyzed for their topology relative to the cytoplasmic or periplasmic side of the membrane and their accessibility to the water space. For the topology studies, DcuB was fused at 33 sites to the reporter enzymes PhoA and LacZ that are only active when located in the periplasm or the cytoplasm, respectively. The ratios of the PhoA and LacZ activities suggested the presence of 10 or 11 hydrophilic loops, and 11 or 12 α-helical transmembrane domains (TMDs). The central part…
Interaction of Heparins and Dextran Sulfates with a Mesoscopic Protein Nanopore
2009
A mechanism of how polyanions influence the channel formed by Staphylococcus aureus alpha-hemolysin is described. We demonstrate that the probability of several types of polyanions to block the ion channel depends on the presence of divalent cations and the polyanion molecular weight and concentration. For heparins, a 10-fold increase in molecular weight decreases the half-maximal inhibitory concentration, IC(50), nearly 10(4)-fold. Dextran sulfates were less effective at blocking the channel. The polyanions are significantly more effective at reducing the conductance when added to the trans side of this channel. Lastly, the effectiveness of heparins on the channel conductance correlated wi…
A Na+-coupled C4-dicarboxylate transporter (Asuc_0304) and aerobic growth of Actinobacillus succinogenes on C4-dicarboxylates
2014
Actinobacillus succinogenes, which is known to produce large amounts of succinate during fermentation of hexoses, was able to grow on C4-dicarboxylates such as fumarate under aerobic and anaerobic conditions. Anaerobic growth on fumarate was stimulated by glycerol and the major product was succinate, indicating the involvement of fumarate respiration similar to succinate production from glucose. The aerobic growth on C4-dicarboxylates and the transport proteins involved were studied. Fumarate was oxidized to acetate. The genome of A. succinogenes encodes six proteins with similarity to secondary C4-dicarboxylate transporters, including transporters of the Dcu (C4-dicarboxylate uptake), Dcu…
Impact of exogenous lactate on survival and radioresponse of carcinoma cells in vitro
2009
Tumour lactate levels have been shown to correlate with high radioresistance in tumour models in vivo. The study aimed to evaluate the impact of pathophysiological extracellular lactate concentrations and acidosis on the in vitro survival and radioresponse of various cancer cell lines.HCT-116, HT29 (colorectal) and FaDu (HNSCC) carcinoma cells were studied. Lactate release rates were determined, and expression of the monocarboxylate transporter MCT1 and its cofactor CD147 were monitored by immunofluorescence and flow cytometry. Colony formation was compared for cells exposed to 20 mM exogenous lactate, acidosis (pH 6.4) and lactate plus acidosis relative to control and dose response curves …