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Taxol, a natural plant product that enhances the rate and extent of microtubule assembly in vitro and stabilizes microtubules in vitro and in cells, was labeled with tritium by catalytic exchange with (3)H(2)O. The binding of [(3)H]taxol to microtubule protein was studied by a sedimentation assay. Microtubules assembled in the presence of [(3)H]taxol bind drug specifically with an apparent binding constant, K(app), of 8.7 x 19(-7) M and binding saturates with a calculated maximal binding ration, B(max), of 0.6 mol taxol bound/mol tubulin dimer. [(3)H]Taxol also binds and assembles phosphocellulose-purified tubulin, and we suggest that taxol stabilizes interactions between dimers that lead to microtubule polymer formation. With both microtubule protein and phosphocellulose- purified tubulin, binding saturation occurs at approximate stoichiometry with the tubulin dimmer concentration. Under assembly conditions, podophyllotoxin and vinblastine inhibit the binding of [(3)H]taxol to microtubule protein in a complex manner which we believe reflects a competition between these drugs, not for a single binding site, but for different forms (dimer and polymer) of tubulin. Steady-state microtubules assembled with GTP or with 5’-guanylyl-α,β-methylene diphosphonate (GPCPP), a GTP analog reported to inhibit microtubule treadmilling (I.V. Sandoval and K. Weber. 1980. J. Biol. Chem. 255:6966-6974), bind [(3)H]taxol with approximately the same stoichiometry as microtubules assembled in the presence of [(3)H]taxol. Such data indicate that a taxol binding site exists on the intact microtubule. Unlabeled taxol competitively displaces [(3)H]taxol from microtubules, while podophyllotoxin, vinblastine, and CaCl(2) do not. Podophyllotoxin and vinblastine, however, reduce the mass of sedimented taxol-stabilized microtubules, but the specific activity of bound [(3)H]taxol in the pellet remains constant. We conclude that taxol binds specifically and reversibly to a polymerized form of tubulin with a stoichiometry approaching unity. 相似文献
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Stephen A Bustin Jean-François Beaulieu Jim Huggett Rolf Jaggi Frederick SB Kibenge Pål A Olsvik Louis C Penning Stefan Toegel 《BMC molecular biology》2010,11(1):74
The conclusions of thousands of peer-reviewed publications rely on data obtained using fluorescence-based quantitative real-time
PCR technology. However, the inadequate reporting of experimental detail, combined with the frequent use of flawed protocols
is leading to the publication of papers that may not be technically appropriate. We take the view that this problem requires
the delineation of a more transparent and comprehensive reporting policy from scientific journals. This editorial aims to
provide practical guidance for the incorporation of absolute minimum standards encompassing the key assay parameters for accurate
design, documentation and reporting of qPCR experiments (MIQE précis) and guidance on the publication of pure 'reference gene'
articles. 相似文献
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Herbert SB Baraf Michael A Becker Sergio R Gutierrez-Urena Edward L Treadwell Janitzia Vazquez-Mellado Claudia D Rehrig Faith D Ottery John S Sundy Robert A Yood 《Arthritis research & therapy》2013,15(5):R137
Introduction
Two replicate randomized, placebo-controlled six-month trials (RCTs) and an open-label treatment extension (OLE) comprised the pegloticase development program in patients with gout refractory to conventional therapy. In the RCTs, approximately 40% of patients treated with the approved dose saw complete response (CR) of at least one tophus. Here we describe the temporal course of tophus resolution, total tophus burden in patients with multiple tophi, tophus size at baseline, and the relationship between tophus response and urate-lowering efficacy.Methods
Baseline subcutaneous tophi were analyzed quantitatively using computer-assisted digital images in patients receiving pegloticase (8 mg biweekly or monthly) or placebo in the RCTs, and pegloticase in the OLE. Tophus response, a secondary endpoint in the trials, was evaluated two ways. Overall tophus CR was the proportion of patients achieving a best response of CR (without any new/enlarging tophi) and target tophus complete response (TT-CR) was the proportion of all tophi with CR.Results
Among 212 patients randomized in the RCTs, 155 (73%) had ≥1 tophus and 547 visible tophi were recorded at baseline. Overall tophus CR was recorded in 45% of patients in the biweekly group (P = 0.002 versus placebo), 26% in the monthly group, and 8% in the placebo group after six months of RCT therapy. TT-CR rates at six months were 28%, 19%, and 2% of tophi, respectively. Patients meeting the primary endpoint of sustained urate-lowering response to therapy (responders) were more likely than nonresponders to have an overall tophus CR at six months (54% vs 20%, respectively and 8% with placebo).Both overall tophus CR and TT-CRs increased with treatment duration in the OLE, reaching 70% (39/56) of patients and 55% (132/238) of target tophi after one year of treatment in patients receiving pegloticase during both the RCTs and OLE. At that time point, more tophi had resolved in responders (102/145 or 70% of tophi) than nonresponders (30/93; 32%).Conclusions
Pegloticase reduced tophus burden in patients with refractory tophaceous gout, especially those achieving sustained urate-lowering. Complete resolution of tophi occurred in some patients by 13 weeks and in others with longer-term therapy.Trial registrations
, NCT00325195 NCT01356498相似文献36.
INTRODUCTIONAsearlyasin1948wehavefr8CtionatedisolatednucleifromnormalandtumorcellsbyextractionwithiMNaCIanddilutealkali[1].Thenuclearresiduewasthenstudiedmorethoroughly[2,3].Lateron,sillillarproteinousnuclearresidueswereisolatedbyotherworkers[46]andasstud… 相似文献
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Post ‘omic’ era has resulted in the development of many primary, secondary and derived databases. Many analytical and
visualization bioinformatics tools have been developed to manage and analyze the data available through large sequencing
projects. Availability of heterogeneous databases and tools make it difficult for researchers to access information from varied
sources and run different bioinformatics tools to get desired analysis done. Building integrated bioinformatics platforms is one of
the most challenging tasks that bioinformatics community is facing. Integration of various databases, tools and algorithm is a
challenging problem to deal with. This article describes the bioinformatics analysis workflow management systems that are
developed in the area of gene sequence analysis and phylogeny. This article will be useful for biotechnologists, molecular
biologists, computer scientists and statisticians engaged in computational biology and bioinformatics research. 相似文献
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Mezghanni H Khedher SB Tounsi S Zouari N 《Preparative biochemistry & biotechnology》2012,42(3):267-278
In order to overproduce biofungicides agents by Bacillus amyloliquefaciens BLB371, a suitable culture medium was optimized using response surface methodology. Plackett-Burman design and central composite design were employed for experimental design and analysis of the results. Peptone, sucrose, and yeast extract were found to significantly influence antifungal activity production and their optimal concentrations were, respectively, 20 g/L, 25 g/L, and 4.5 g/L. The corresponding biofungicide production was 250 AU/mL, corresponding to 56% improvement in antifungal components production over a previously used medium (160 AU/mL). Moreover, our results indicated that a deficiency of the minerals CuSO(4), FeCl(3) · 6H(2)O, Na(2)MoO(4), KI, ZnSO(4) · 7H(2)O, H(3)BO(3), and C(6)H(8)O(7) in the optimized culture medium was not crucial for biofungicides production by Bacillus amyloliquefaciens BLB371, which is interesting from a practical point of view, particularly for low-cost production and use of the biofungicide for the control of agricultural fungal pests. 相似文献
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Intensity normalization is an important pre-processing step in the study and analysis of DaTSCAN SPECT imaging. As most automatic supervised image segmentation and classification methods base their assumptions regarding the intensity distributions on a standardized intensity range, intensity normalization takes on a very significant role. In this work, a comparison between different novel intensity normalization methods is presented. These proposed methodologies are based on Gaussian Mixture Model (GMM) image filtering and mean-squared error (MSE) optimization. The GMM-based image filtering method is achieved according to a probability threshold that removes the clusters whose likelihood are negligible in the non-specific regions. The MSE optimization method consists of a linear transformation that is obtained by minimizing the MSE in the non-specific region between the intensity normalized image and the template. The proposed intensity normalization methods are compared to: i) a standard approach based on the specific-to-non-specific binding ratio that is widely used, and ii) a linear approach based on the α-stable distribution. This comparison is performed on a DaTSCAN image database comprising analysis and classification stages for the development of a computer aided diagnosis (CAD) system for Parkinsonian syndrome (PS) detection. In addition, these proposed methods correct spatially varying artifacts that modulate the intensity of the images. Finally, using the leave-one-out cross-validation technique over these two approaches, the system achieves results up to a 92.91% of accuracy, 94.64% of sensitivity and 92.65 % of specificity, outperforming previous approaches based on a standard and a linear approach, which are used as a reference. The use of advanced intensity normalization techniques, such as the GMM-based image filtering and the MSE optimization improves the diagnosis of PS. 相似文献
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