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201.
Sonia Del Prete Viviana De Luca Daniela Vullo Sameh M. Osman Zeid AlOthman Vincenzo Carginale 《Journal of enzyme inhibition and medicinal chemistry》2016,31(6):1156-1161
Malassezia yeasts are almost exclusively the single eukaryotic members of the fungal flora of the skin. Malassezia globosa and Malassezia restricta are found on the skin of practically all humans. Malassezia globosa is highly implicated in the pathogenesis of dandruff and its genome encodes for only one carbonic anhydrases (CAs, EC 4.2.1.1) belonging to the β-class (MgCA). It has been indeed demonstrated that in many pathogenic microorganisms, CAs are essential for their life cycle and their inhibition can lead to growth impairment and defects. In the previous work, the recombinant MgCA was investigated for its inhibition profile with sulfonamides, which in models of dandruff infection were able to protect animals from the fungal infection, allowing us to propose this enzyme as a new antidandruff target. MgCA was cloned as GST-fusion protein, but the yield was rather low and the protein was often found in inclusion bodies. Here, we propose an alternative procedure consisting in cloning the recombinant MgCA as His-Tag fusion protein. This procedure resulted in a good method to express and purify the active recombinant MgCA, and the protein recovery was better with respect to that used for preparing MG-CA (β-CA cloned as GST-fusion protein). 相似文献
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Alessio Giacomini Viviana Corich Francisco J. Ollero rea Squartini Marco P. Nuti 《FEMS microbiology letters》1992,100(1-3):87-90
Several experimental conditions and parameters contributing to the determination of beta-galactosidase activity, as proposed in Miller's assay, were studied. Use of the absorbance correction factor and the nature and concentration of permeabilizing agents were taken into account as different experimental conditions. Reaction time, culture volume, and growth stage were investigated as equation parameters. From a quantitative point of view the results, in terms of Miller units, are markedly affected by variation in these conditions. Therefore, to ensure reproducibility it is advisable to use constant values for all the parameters. 相似文献
205.
Llop E Gallego RG Belalcazar V Gerwig GJ Kamerling JP Segura J Pascual JA 《Proteomics》2007,7(23):4278-4291
The structure, function, and physico-chemical properties of many proteins are determined by PTM, being glycosylation the most complex. This study describes how a combination of typical proteomics methods (2-DE) combines with glycomics strategies (HPLC, MALDI-TOF-MS, exoglycosidases sequencing) to yield comprehensive data about single spot-microheterogeneity, providing meaningful information for the detection of disease markers, pharmaceutical industry, antidoping control, etc. Recombinant erythropoietin and its hyperglycosylated analogue darbepoetin-alpha were chosen as showcases because of their relevance in these fields and the analytical challenge they represent. The combined approach yielded good results in terms of sample complexity (mixture glycoforms), reproducibility, sensitivity ( approximately 25 pmoles of glycoprotein/spot), and identification of the underlying protein. Heterogeneity was present in all spots but with a clear tendency; spots proximal to the anode contained the highest amount of tetra-antennary tetra-sialylated glycans, whereas the opposite occurred for spots proximal to the cathode with the majority of the structures being undersialylated. Spot microheterogeneity proved a consequence of the multiple glycosylation sites as they contributed directly to the number of possibilities to account for a discrete charge in a single spot. The interest of this combined glycoproteomics method resides in the efficiency for detecting and quantifying subtle dissimilarities originated from altered ratios of identical glycans including N-acetyl-lactosamine repeats, acetylation, or antigenic epitopes, that do not significantly contribute to the electrophoretic mobility, but affect the glycan microheterogeneity and the potential underlying related functionality. 相似文献
206.
Type III secretion systems of Gram-negative pathogenic bacteria allow the injection of effector proteins into the cytosol of host eukaryotic cells. Crossing of the eukaryotic plasma membrane is facilitated by a translocon, an oligomeric structure made up of two bacterial proteins inserted into the host membrane during infection. In Pseudomonas aeruginosa, a major human opportunistic pathogen, these proteins are PopB and PopD. Their interactions with their common chaperone PcrH in the cytosol of the bacteria are essential for the proper function of the injection system. The interaction region between PopD and PcrH was identified using limited proteolysis, revealing that the putative PopD transmembrane fragment is buried within the PopD/PcrH complex. In addition, structural features of PopD and PcrH, either individually or within the binary complex, were characterized using spectroscopic methods and 1D NMR. Whereas PcrH possesses the characteristics of a folded protein, PopD is in a molten globule state either alone or in the PopD/PcrH complex. The molten globule state is known to enable the membrane insertion of translocation/pore-forming domains of bacterial toxins. Therefore, within the bacterial cytoplasm, PopD is preserved in a state that is favorable to secretion and insertion into cell membranes. 相似文献
207.
Norma Candia Beatriz Lopez Thierry Zozio Marcela Carrivale Chyntia Diaz Graciela Russomando Nilda J de Romero Juan C Jara Lucia Barrera Nalin Rastogi Viviana Ritacco 《BMC microbiology》2007,7(1):1-10
Background
Francisella tularensis is a gram negative, facultative intracellular bacterium that is the etiological agent of tularemia. F. novicida is closely related to F. tularensis but has low virulence for humans while being highly virulent in mice. IglA is a 21 kDa protein encoded by a gene that is part of an iglABCD operon located on the Francisella pathogenicity island (FPI).Results
Bioinformatics analysis of the FPI suggests that IglA and IglB are components of a newly described type VI secretion system. In this study, we showed that IglA regulation is controlled by the global regulators MglA and MglB. During intracellular growth IglA production reaches a maximum at about 10 hours post infection. Biochemical fractionation showed that IglA is a soluble cytoplasmic protein and immunoprecipitation experiments demonstrate that it interacts with the downstream-encoded IglB. When the iglB gene was disrupted IglA could not be detected in cell extracts of F. novicida, although IglC could be detected. We further demonstrated that IglA is needed for intracellular growth of F. novicida. A non-polar iglA deletion mutant was defective for growth in mouse macrophage-like cells, and in cis complementation largely restored the wild type macrophage growth phenotype.Conclusion
The results of this study demonstrate that IglA and IglB are interacting cytoplasmic proteins that are required for intramacrophage growth. The significance of the interaction may be to secrete effector molecules that affect host cell processes. 相似文献208.
Dendritic cells are the major antigen-presenting and antigen-priming cells of the immune system. We review the antigen-presenting
and proinflammatory roles played by dendritic cells in the initiation of rheumatoid arthritis (RA) and atherosclerosis, which
complicates RA. Various signals that promote the activation of NF-κB and the secretion of TNF and IL-1 drive the maturation
of dendritic cells to prime self-specific responses, and drive the perpetuation of synovial inflammation. These signals may
include genetic factors, infection, cigarette smoking, immunostimulatory DNA and oxidized low-density lipoprotein, with major
involvement of autoantibodies. We propose that the pathogenesis of RA and atherosclerosis is intimately linked, with the vascular
disease of RA driven by similar and simultaneous triggers to NF-κB. 相似文献
209.
Gastroschisis and young mothers: What makes them different from other mothers of the same age? 下载免费PDF全文
210.