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991.
Feng J Chen Y Pu J Yang X Zhang C Zhu S Zhao Y Yuan Y Yuan H Liao F 《Analytical biochemistry》2011,(1):144-149
The classical malachite green (MLG) assay of phosphate, which added MLG after molybdate to the acidified reaction solutions of phosphate, tolerated interference from papaverine, sildenafil, and some similar hydrophobic amines. Resonance Rayleigh scattering signals, the alleviation of interference by poly(vinyl alcohol), and the precipitation of some yellow complexes supported that the irreversible aggregation of the complexes of a hydrophobic amine of interference and phosphomolybdate reduced the amounts of phosphomolybdate accessible to MLG and caused the interference. By adding MLG before molybdate to the acidified reaction solutions of phosphate, the complexes of phosphomolybdate and MLG were preferentially formed before the complexes of phosphomolybdate and such a hydrophobic amine effectively aggregated; thereby, an improved MLG assay of phosphate with the resistance to common hydrophobic amines was developed. Using the improved MLG assay of phosphate and a phosphatase to release phosphate from AMP, a spectrometric method successfully estimated the half-inhibition concentrations of papaverine on the recombinant human cyclic nucleotide phosphodiesterase (PDE) isozyme 4 and the mixture of PDE isozymes from rabbit brain. Therefore, the improved MLG assay of phosphate was a favorable and universal technique for developing spectrometric methods for characterizing and screening inhibitors of enzymes that release phosphate during their actions. 相似文献
992.
The reaction mechanism of mycolic acid cyclopropane synthase is investigated using hybrid density functional theory. The direct methylation mechanism is examined with a large model of the active site constructed on the basis of the crystal structure of the native enzyme. The important active site residue Glu140 is modeled in both ionized and neutral forms. We demonstrate that the reaction starts via the transfer of a methyl to the substrate double bond, followed by the transfer of a proton from the methyl cation to the bicarbonate present in the active site. The first step is calculated to be rate-limiting, in agreement with experimental kinetic results. The protonation state of Glu140 has a rather weak influence on the reaction energetics. In addition to the natural reaction, a possible side reaction, namely a carbocation rearrangement, is also considered and is shown to have a low barrier. Finally, the energetics for the sulfur ylide proposal, which has already been ruled out, is also estimated, showing a large energetic penalty for ylide formation. 相似文献
993.
Bacterial cells evolved under prolonged stress often have a growth advantage in stationary phase (GASP); we expect GASP cells to maintain a proliferative state and dominate wild-type cells during starvation, especially when nutrients are limited and the medium has been conditioned. However, when we compete GASP mutants against wild-type cells in a chain of microfluidic microhabitat patches (MHPs) with alternating nutrient-rich and nutrient-limited regions, we observe the reverse effect: wild-type cells achieve maximum relative density under nutrient-limited conditions, while GASP cells dominate nutrient-rich regions. We explain this surprising observation in terms of ideal free distributions, where we show that wild-type cells maximize their fitness at high cell density by redistributing themselves to sparsely populated MHPs. At the microscopic level, we describe how biofilm formation also contributes to the population redistribution. We conclude by discussing the implications of these results for social interactions of more complex organisms. 相似文献
994.
Liao YC Huang TW Chen FC Charusanti P Hong JS Chang HY Tsai SF Palsson BO Hsiung CA 《Journal of bacteriology》2011,193(7):1710-1717
Klebsiella pneumoniae is a Gram-negative bacterium of the family Enterobacteriaceae that possesses diverse metabolic capabilities: many strains are leading causes of hospital-acquired infections that are often refractory to multiple antibiotics, yet other strains are metabolically engineered and used for production of commercially valuable chemicals. To study its metabolism, we constructed a genome-scale metabolic model (iYL1228) for strain MGH 78578, experimentally determined its biomass composition, experimentally determined its ability to grow on a broad range of carbon, nitrogen, phosphorus and sulfur sources, and assessed the ability of the model to accurately simulate growth versus no growth on these substrates. The model contains 1,228 genes encoding 1,188 enzymes that catalyze 1,970 reactions and accurately simulates growth on 84% of the substrates tested. Furthermore, quantitative comparison of growth rates between the model and experimental data for nine of the substrates also showed good agreement. The genome-scale metabolic reconstruction for K. pneumoniae presented here thus provides an experimentally validated in silico platform for further studies of this important industrial and biomedical organism. 相似文献
995.
T cell immunoglobulin and mucin domain (Tim)-3 is expressed on activated CD4+ and CD8+ T cells. Identification of galectin-9 as a ligand for Tim-3 has now firmly established the Tim-3/galectin-9 pathway, which results in apoptosis of effector CD4+ and CD8+ T cells. Moreover, Th17 cells are a recently discovered CD4+ effector T cell, which are important in antimicrobial immunity. Whether the Tim-3/galectin-9 pathway affects Th17 immunity has not been elucidated. Here, we demonstrated expression of Tim-3 on Th17 cells by flow cytometry. Th17-skewed cells were sensitive to galectin-9-induced apoptosis. In vitro administration of galectin-9 decreased stimulated Th17 cells and inhibited production of IL-17. Interestingly, Klebsiella pneumoniae (K. pneumoniae) infection led to enhanced IL-17 levels. Recombinant galectin-9 significantly decreased IL-17 in vivo, which resulted in reduced bacterial clearance and high mortality. These observations suggest that the Tim-3/galectin-9 pathway plays an important role in termination of Th17-immune responses, and could be a therapeutic target for inflammatory diseases. 相似文献
996.
Aims
Some rhizobia can convert insoluble P into available forms for plant growth but the underlying mechanisms for this are not understood. In this study, the function of rhizobia in P acquisition from P sources for soybean was studied.Methods
Four rhizobial strains were employed to evaluate their phosphate-solubilizing (PS) activity, their ability to mediate pH changes in growth medium for different P sources, and IAA production. A sand culture experiment using different P sources was carried out to characterize P acquisition changes of soybean plants with or without rhizobium inoculation. Rhizospheric acidification in soybean was further analyzed in hydroponics.Results
Our results showed that all the tested rhizobial strains exhibited significant PS activity for different P sources in the order of Ca-P>Al-P>Phy-P??Fe-P as indicated by the halo/colony ratio technique and increased Pi percentage in the solid and liquid phases, respectively. Furthermore, all of the rhizobial strains could acidify the growth medium for all P sources except Phy-P, but only three of them produced IAA. Compared to non-nodulated plants, the nodulated plants had greater plant biomass and P content in sand culture for all the tested P sources, especially for Ca-P. Moreover, H+ and total acid exudation was more significantly enhanced in the nodulated plants in hydroponics.Conclusions
Our results suggested that the PS ability of rhizobia is more related to acidification of the growth medium than IAA production. Rhizobium inoculation could enhance P acquisition in soybean, especially on soils where Ca-P is the primary P source, and the primary mechanism for rhizobial-mediated P solubilization appears to be via Pi remobilization of nodulated roots through rhizospheric acidification. 相似文献997.
Late embryogenesis abundant (LEA) group 4 (LEA4) proteins play an important role in the water stress tolerance of plants. Although they have been hypothesized to stabilize macromolecules in stressed cells, the protective functions and mechanisms of LEA4 proteins are still not clear. In this study, the metal binding properties of two related soybean LEA4 proteins, GmPM1 and GmPM9, were tested using immobilized metal ion affinity chromatography (IMAC). The metal ions Fe(3+), Ni(2+), Cu(2+) and Zn(2+) were observed to bind these two proteins, while Ca(2+), Mg(2+) or Mn(2+) did not. Results from isothermal titration calorimetry (ITC) indicated that the binding affinity of GmPM1 for Fe(3+) was stronger than that of GmPM9. Hydroxyl radicals generated by the Fe(3+)/H(2)O(2) system were scavenged by both GmPM1 and GmPM9 in the absence or the presence of high ionic conditions (100 mM NaCl), although the scavenging activity of GmPM1 was significantly greater than that of GmPM9. These results suggest that GmPM1 and GmPM9 are metal-binding proteins which may function in reducing oxidative damage induced by abiotic stress in plants. 相似文献
998.
999.
Dan Wang Jing Mao Bin Zhou Xiao-Fu Liao Shi-Qiang Gong Yan Liu Jing-Tao Zhang 《Biotechnology letters》2011,33(1):191-197
Purpose of work
Our study provides a promising alternative of biomimetic coating which functionalizes the dental implant with adhesion peptides and may be useful for enhancing the bone remodeling around Ti implants. 相似文献1000.
Hyoscyamine 6β-hydroxylase (H6H; EC 1.14.11.11) converts hyoscyamine to scopolamine in the last step of scopolamine biosynthetic
pathway. The gene encoding H6H in Anisodus acutangulus was cloned and expressed in Escherichia coli and the recombinant proteins fused with His-tag or GST-tag at its N-terminal were purified and then confirmed by Western bolt analysis. The biofunctional assay revealed that the His-AaH6H and
GST-AaH6H converted hyoscyamine (40 mg/l) to scopolamine at 32 and 31 mg/l, respectively. This is the first report on AaH6H
expression, purification and functional characterization facilitates further genetic improvement of scopolamine yield in A. acutangulus. 相似文献