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1.
《Journal of molecular biology》2021,433(19):167162
Many proteins that can assemble into higher order structures termed amyloids can also concentrate into cytoplasmic inclusions via liquid–liquid phase separation. Here, we study the assembly of human Golgi-Associated plant Pathogenesis Related protein 1 (GAPR-1), an amyloidogenic protein of the Cysteine-rich secretory proteins, Antigen 5, and Pathogenesis-related 1 proteins (CAP) protein superfamily, into cytosolic inclusions in Saccharomyces cerevisiae. Overexpression of GAPR-1-GFP results in the formation GAPR-1 oligomers and fluorescent inclusions in yeast cytosol. These cytosolic inclusions are dynamic and reversible organelles that gradually increase during time of overexpression and decrease after promoter shut-off. Inclusion formation is, however, a regulated process that is influenced by factors other than protein expression levels. We identified N-myristoylation of GAPR-1 as an important determinant at early stages of inclusion formation. In addition, mutations in the conserved metal-binding site (His54 and His103) enhanced inclusion formation, suggesting that these residues prevent uncontrolled protein sequestration. In agreement with this, we find that addition of Zn2+ metal ions enhances inclusion formation. Furthermore, Zn2+ reduces GAPR-1 protein degradation, which indicates stabilization of GAPR-1 in inclusions. We propose that the properties underlying both the amyloidogenic properties and the reversible sequestration of GAPR-1 into inclusions play a role in the biological function of GAPR-1 and other CAP family members. 相似文献
2.
A. Klockow-Beck A. Nick St. Geisshuesler D. Schaufelberger 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》1998,720(1-2):141-151
A capillary electrophoresis (CE) method has been developed as an alternative method for the determination of the inorganic degradation products sulfate and sulfamate in topiramate drug product and drug substance, currently performed by ion chromatography. The anions are separated in a background electrolyte containing potassium chromate and boric acid, followed by indirect UV detection. By adding tetradecyltrimethylammonium bromide to the electrolyte, analysis is performed under co-electroosmotic flow conditions. Variations in injection volumes and migration times are compensated for by use of an internal standard. The validation of the method, which was performed according to ICH guidelines (International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use) [1], comprises specificity, accuracy, linearity, precision, sensitivity and robustness. In addition, the results of an actual tablet sample analysis obtained by this CE method are statistically shown to be in close agreement with those obtained by an ion chromatographic method. 相似文献
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4.
Georg Steinhauser Johannes H. Sterba Karin Poljanc Max Bichler Karl Buchtela 《Journal of trace elements in medicine and biology》2006,20(3):119-153
In this study, 18 partly commercially available samples of rock salt from Austria, Germany, Pakistan, Poland, Switzerland, and Ukraine were investigated with respect to their content of trace elements using instrumental neutron activation analysis. Elements detected were Al, Ba, Br, Ca, Ce, Cl, Co, Cr, Cs, Eu, Fe, Hf, La, Mn, Na, Rb, Sb, Sc, Sm, Sr, Ta, Tb, Th, and Zn, some of them only in individual cases. An estimation of the bioavailability of these trace elements was performed by dissolving an equivalent of the sodium chloride samples in diluted hydrochloric acid (simulating stomach acid), filtering off the insoluble components, and analyzing the evaporated filtrate. It could be shown that in most cases bioactive trace elements like Fe can be found in rock salt in the form of almost insoluble compounds and are therefore not significantly bioavailable, whereas thorium, for example, was partly bioavailable in two cases. A significant contribution to the recommended daily intake of metal trace elements by using rock salt for nutrition can be excluded. 相似文献
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The phosphorylation of the proteins of the erythrocyte membrane of patients suffering from hereditary spherocytosis is investigated in intact erythrocytes by their incubation in the presence of radioactive inorganic phosphate. Examination of the phosphorylated components by high-resolution two-dimensional gel electrophoresis reveals only one defect in the pathological membranes, a depressed phosphorylation of the smaller polypeptide of spectrin; band 2. The phosphorylation of band 2 is measured with reference to the phosphorylation of syndein (). In patients showing overt clinical symptoms and for whom splenectomy is advocated the phosphorylation of band 2 is depressed by approx. 70%. After splenectomy the phosphorylation of membrane proteins is restored to normal levels. 相似文献
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L. Goldman 《European biophysics journal : EBJ》1989,16(6):321-325
The time constant of the process producing the delay in Na inactivation development as determined by the two pulse method (delay) was extracted and compared to that of the slowest Na activation process 3 for the I
Na during the conditioning pulse of that same determination. delay and two pulse inactivation c values were computer generated using a nonlinear least squares algorithm. h and single pulse inactivation h values were independently generated for each determination also with the aid of the computer using the same non-linear least squares algorithm. In one determination at 2 mV, c was 4.68 and delay 0.494 ms while h was 4.70 and 3 0.491 ms for a c/h of 0.996 and a delay/3 of 1.006. Mean delay/3 from five determinations in four axons, both Cs and K perfused, and spanning a potential range of-27 to 2mV was 1.068. The precursor process to inactivation is channel opening. Some fraction of channels presumably inactivate via another route where prior channel opening is not required. 相似文献
9.
Rat ovarian membrane LH/hCG receptor was solubilized in various detergents and reconstituted into proteoliposomes. Upon removal of sodium cholate by active absorption on Bio-Beads SM-2, the functional interaction between receptor and adenylate cyclase was restored. Adenylate cyclase was stimulated by hCG, HCG+GTP or GppNHp and NaF. Reconstituted proteoliposomes bound more 125I-hCG (528 fmol/mg protein) than membrane-bound receptors (384 fmol/mg protein). There was no difference, however, in the relative affinity of reconstituted receptor preparations for hCG. 相似文献
10.
The ethanol-oxidizing, proton-reducing Pelobacter acetylenicus was grown in chemostat cocultures with either Acetobacterium woodii, Methanobacterium bryantii, or Desulfovibrio desulfuricans. Stable steady state conditions with tightly coupled growth were reached at various dilution rates between 0.02 and 0.14 h-1. Both ethanol and H2 steady state concentrations increased with growth rate and were lower in cocultures with the sulfate reducer < methanogen < homoacetogen. Due to the higher affinity for H2, D. desulfuricans outcompeted M. bryantii, and this one A. woodii when inoculated in cocultures with P. acetylenicus. Cocultures with A. woodii had lower H2 steady state concentrations when bicarbonate reduction was replaced by the energetically more favourable caffeate reduction. Similarly, cocultures with D. desulfuricans had lower H2 concentrations with nitrate than with sulfate as electron acceptor. The Gibbs free energy (G) available to the H2-producing P. acetylenicus was independent of growth rate and the H2-utilizing partner, whereas the G available to the latter increased with growth rate and the energy yielding potential of the H2 oxidation reaction. The critical Gibbs free energy (Gc), i.e. the minimum energy required for H2 production and H2 oxidation, was-5.5 to-8.0 kJ mol-1 H2 for P. acetylenicus,-5.1 to-6.3 kJ mol-1 H2 for A. woodii,-7.5 to-9.1 kJ mol-1 H2 for M. bryantii, and-10.3 to-12.3 kJ mol-1 H2 for D. desulfuricans. Obviously, the potentially available energy was used more efficiently by homoacetogens > methanogens > sulfate reducers. 相似文献