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1.
Recombinant maltose-binding protein from Thermotoga maritima (TmMBP) was expressed in Escherichia coli and purified to homogeneity, applying heat incubation of the crude extract at 75 degrees C. As taken from the spectral, physicochemical and binding properties, the recombinant protein is indistinguishable from the natural protein isolated from the periplasm of Thermotoga maritima. At neutral pH, TmMBP exhibits extremely high intrinsic stability with a thermal transition >105 degrees C. Guanidinium chloride-induced equilibrium unfolding transitions at varying temperatures result in a stability maximum at approximately 40 degrees C. At room temperature, the thermodynamic analysis of the highly cooperative unfolding equilibrium transition yields DeltaG(N-->U)=100(+/-5) kJ mol(-1 )for the free energy of stabilization. Compared to mesophilic MBP from E. coli as a reference, this value is increased by about 60 kJ mol(-1). At temperatures around the optimal growth temperature of T. maritima (t(opt) approximately 80 degrees C), the yield of refolding does not exceed 80 %; the residual 20 % are misfolded, as indicated by a decrease in stability as well as loss of the maltose-binding capacity. TmMBP is able to bind maltose, maltotriose and trehalose with dissociation constants in the nanomolar to micromolar range, combining the substrate specificities of the homologs from the mesophilic bacterium E. coli and the hyperthermophilic archaeon Thermococcus litoralis. Fluorescence quench experiments allowed the dissociation constants of ligand binding to be quantified. Binding of maltose was found to be endothermic and entropy-driven, with DeltaH(b)=+47 kJ mol(-1) and DeltaS(b)=+257 J mol(-1) K(-1). Extrapolation of the linear vant'Hoff plot to t(opt) resulted in K(d) approximately 0.3 microM. This result is in agreement with data reported for the MBPs from E. coli and T. litoralis at their respective optimum growth temperatures, corroborating the general observation that proteins under their specific physiological conditions are in corresponding states. 相似文献
2.
K Berr D Wassenberg H Lilie J Behlke R Jaenicke 《European journal of biochemistry》2000,267(17):5413-5420
Taxon-specific epsilon-crystallin (epsilonC) from duck eye lens is identical to duck heart muscle lactate dehydrogenase. It forms a dimer of dimers with a dissociation constant of 2.2 x 10-7 M, far beyond the value observed for other vertebrate lactate dehydrogenases. Comparing the characteristics of wild-type epsilon-crystallin with those of three mutants, G115N, G119F and 115N/119F, representing the only significant peripheral sequence variations between duck epsilonC and chicken or pig heart muscle lactate dehydrogenase, no significant conformational differences are detectable. Regarding the catalytic properties, the Michaelis constant of the double mutant 115N/119F for pyruvate is found to be decreased; for wild-type enzyme, the effect is overcompensated by the high expression level of epsilonC in the eye lens. As taken from spectral analysis of the guanidine-induced and temperature-induced denaturation transitions, epsilonC in its dimeric state is relatively unstable, whereas the native tetramer exhibits the high intrinsic stability characteristic of common vertebrate heart and muscle lactate dehydrogenases. The denaturation mechanism of epsilonC is complex and only partially reversible. In the case of thermal unfolding, the predominant side reaction competing with the reconstitution of the native state is the kinetic partitioning between proper folding and aggregation. alpha-Crystallin, the major molecular chaperone in the eye lens, inhibits the aggregation of epsilonC by trapping the misfolded protein. 相似文献
3.
van der Heijde D Landewé R Boonen A Einstein S Herborn G Rau R Wassenberg S Weissman BN Winalski CS Sharp JT 《Arthritis research & therapy》2007,9(4):R62
The objective of the present study was to test the hypothesis that experts recognize repair of erosions and, if so, to determine
which, if any, morphologic features permitted them to recognize the repair. We also tested whether scoring by a standard method
detected repair. Seven experienced readers of radiographs in rheumatoid arthritis were presented with 64 sets of single joints-of-interest
at two time points, randomized and blinded for the correct sequence. The readers assessed which joint was better, and recorded
whether any of six specific features were seen. Two independent readers, experienced in scoring by the van der Heijde-modified
Sharp method who were not on the expert panel, then scored the complete films that included the joint-of-interest. The panel
agreed very well on which of two joints was better, and, even though they did not know the true sequence, the panel accurately
assigned a sequence slightly better than chance alone (58%) but worse than their agreement on which image was 'better or worse'
(78%). The readers therefore indirectly assigned repair by choosing the second film as the best. Putative repair features
were seen in cases of both repair and progression, and were not discriminatory. Similar results were obtained when the experts
were presented with the entire hand or foot containing the joint-of-interest. In the third repair exercise, two independent
readers who scored whole hands and feet using a standard method found a mean negative score in 22/60 joints-of-interest. All
22 joints were also scored as repair by the panel. Repair was detected reliably by a majority of the panel on viewing paired
images based on a better/worse decision and assigning sequence in a set of images that were blinded for sequence by an independent
project manager. In this test set of images, repair was manifested by a reduction in the size of erosion in many cases. Size
was one feature that aided the experts to detect repair but cannot be the only one; the experts had to find other features
to determine whether a smaller erosion was the first in a sequence of radiographs in a patient with progressive damage or
was the second film in a patient exhibiting repair. The change in size of erosion was also picked up by independent readers
applying the van der Heijde-modified Sharp scoring method and was reflected in their scores. 相似文献
4.
Abstract. 1. Foragers of social insects can be guided to profitable food sources by social information transfer within the nest. This study showed that in addition to such an information-centre strategy, social information in the field also plays an important role in individual foraging decisions. The effect of the presence of a nestmate on individual decision-making on where to forage was investigated in six species of stingless bee that differ in their recruitment system. Some species preferred to feed close to a nestmate (local enhancement) whereas other species actively avoided landing close to a nestmate. The term local inhibition is introduced for this avoidance behaviour.
2. Local enhancement and local inhibition were species specific but were not related to the species' recruitment system.
3. Local enhancement and local inhibition were affected by the individual's experience with the food source. Newly recruited foragers of Trigona amalthea showed local enhancement whereas experienced foragers showed local inhibition.
4. These individual decision-making rules explained accurately the spatial distribution of recruited nestmates: foraging groups of T. amalthea , which shows local inhibition, were more dispersed than foraging groups of Oxytrigona mellicolor , which shows local enhancement.
5. The effect of heterospecifics on stingless bee flower choice was investigated for 18 species combinations. Landing decisions were influenced significantly by the aggressiveness and the body size of the resident bee. Larger and more aggressive heterospecifics were avoided, whereas in some cases less aggressive bees acted as an attraction cue. 相似文献
2. Local enhancement and local inhibition were species specific but were not related to the species' recruitment system.
3. Local enhancement and local inhibition were affected by the individual's experience with the food source. Newly recruited foragers of Trigona amalthea showed local enhancement whereas experienced foragers showed local inhibition.
4. These individual decision-making rules explained accurately the spatial distribution of recruited nestmates: foraging groups of T. amalthea , which shows local inhibition, were more dispersed than foraging groups of Oxytrigona mellicolor , which shows local enhancement.
5. The effect of heterospecifics on stingless bee flower choice was investigated for 18 species combinations. Landing decisions were influenced significantly by the aggressiveness and the body size of the resident bee. Larger and more aggressive heterospecifics were avoided, whereas in some cases less aggressive bees acted as an attraction cue. 相似文献
5.
Background
The costs and benefits of controlling nosocomial spread of antibiotic-resistant bacteria are unknown.Methods
We developed a mathematical algorithm to determine cost-effectiveness of infection control programs and explored the dynamical interactions between different epidemiological variables and cost-effectiveness. The algorithm includes occurrence of nosocomial infections, attributable mortality, costs and efficacy of infection control and how antibiotic-resistant bacteria affect total number of infections: do infections with antibiotic-resistant bacteria replace infections caused by susceptible bacteria (replacement scenario) or occur in addition to them (addition scenario). Methicillin-resistant Staphylococcus aureus (MRSA) bacteremia was used for illustration using observational data on S. aureus bacteremia (SAB) in our hospital (n = 189 between 2001–2004, all being methicillin-susceptible S. aureus [MSSA]).Results
In the replacement scenario, the costs per life year gained range from € 45,912 to € 6590 for attributable mortality rates ranging from 10% to 50%. Using € 20,000 per life year gained as a threshold, completely preventing MRSA would be cost-effective in the replacement scenario if attributable mortality of MRSA is ≥21%. In the addition scenario, infection control would be cost saving along the entire range of estimates for attributable mortality.Conclusions
Cost-effectiveness of controlling antibiotic-resistant bacteria is highly sensitive to the interaction between infections caused by resistant and susceptible bacteria (addition or replacement) and attributable mortality. In our setting, controlling MRSA would be cost saving for the addition scenario but would not be cost-effective in the replacement scenario if attributable mortality would be <21%. 相似文献6.
Joerg Wendler Gerd R Burmester Helmut S?rensen Andreas Krause Constanze Richter Hans-Peter Tony Andrea Rubbert-Roth Peter Bartz-Bazzanella Siegfried Wassenberg Iris Haug-Rost Thomas D?rner 《Arthritis research & therapy》2014,16(2):R80
Introduction
The aim of this study was to evaluate the safety and efficacy of rituximab (RTX) in a large cohort of patients with rheumatoid arthritis in routine care, and to monitor changes in daily practice since the introduction of RTX therapy.Methods
This was a multicentre, prospective, non-interventional study conducted under routine practice conditions in Germany. Efficacy was evaluated using Disease Activity Score in 28 joints (DAS28) and Health Assessment Questionnaire-Disability Index (HAQ-DI). Safety was assessed by recording adverse drug reactions (ADRs). Physician and patient global efficacy and tolerability assessments were also evaluated.Results
Overall, 2,484 patients (76.7% female, mean age 56.4 years, mean disease duration 11.7 years) received RTX treatment (22.7% monotherapy). The total observation period was approximately six-years (median follow-up 14.7 months). RTX treatment led to improvements in DAS28 and HAQ-DI that were sustained over multiple courses. DAS28 improvements positively correlated with higher rheumatoid factor levels up to 50 IU/ml. Response and tolerability were rated good/very good by the majority of physicians and patients. Mean treatment intervals were 10.5 and 6.8 months for the first and last 400 enrolled patients, respectively. Infections were the most frequently reported ADRs (9.1%; 11.39/100 patient-years); approximately 1% of patients per course discontinued therapy due to ADRs.Conclusions
Prolonged RTX treatment in routine care is associated with good efficacy and tolerability, as measured by conventional parameters and by physicians’ and patients’ global assessments. Rheumatoid factor status served as a distinct and quantitative biomarker of RTX responsiveness. With growing experience, physicians repeated treatments earlier in patients with less severe disease activity. 相似文献7.
8.
Thermotoga maritima XynA is an extremely thermostable modular enzyme with five domains (A1-A2-B-C1-C2). Its catalytic domain (-B-) is flanked by duplicated non-catalytic domains. The C-terminal repeated domains represent cellulose-binding domains (CBDs). Xylanase domains related to the N-terminal domains of XynA (A1-A2) are called thermostabilizing domains because their deletion normally leads to increased thermosensitivity of the enzymes. It was found that a glutathione-S-transferase (GST) hybrid protein (GST-A1A2) containing both A-domains of XynA can interact with various soluble xylan preparations and with mixed-linkage beta-1,3/beta-1,4-glucans. GST-A1A2 showed no affinity for insoluble microcrystalline cellulose, whereas, vice versa, GST-C2, which contains the C-terminal CBD of XynA, did not interact with soluble xylan. Another hybrid protein, GST-A2, displayed the same binding properties as GST-A1A2, indicating that A2 alone can also promote xylan binding. The dissociation constants for the binding of xylose, xylobiose, xylotriose, xylotetraose and xylopentaose by GST-A2, as determined at 20 degrees C by fluorescence quench experiments, were 8.1 x 10(-3) M, 2.3 x 10(-4) M, 2.3 x 10(-5) M, 2.5 x 10(-6)M and 1.1 x 10(-6) M respectively. The A-domains of XynA, which are designated as xylan binding domains (XBD), are, from the structural as well as the functional point of view, prototypes of a novel class of binding domains. More than 50 related protein segments with hitherto unknown function were detected in about 30 other multidomain beta-glycanases, among them putative plant (Arabidopsis thaliana) xylanases. It is argued that polysaccharide binding and not thermostabilization is the main function of A-like domains. 相似文献
9.
Background
Preoperative screening for nasal S. aureus carriage, followed by eradication treatment of identified carriers with nasal mupirocine ointment and chlorhexidine soap was highly effective in preventing deep-seated S. aureus infections. It is unknown how cost-effectiveness of this intervention is affected by suboptimal S. aureus screening. We determined cost-effectiveness of different preoperative S. aureus screening regimes.Methods
We compared different screening scenarios (ranging from treating all patients without screening to treating only identified S. aureus carriers) to the base case scenario without any screening and treatment. Screening and treatment costs as well as costs and mortality due to deep-seated S. aureus infection were derived from hospital databases and prospectively collected data, respectively.Results
As compared to the base case scenario, all scenarios are associated with improved health care outcomes at reduced costs. Treating all patients without screening is most cost-beneficial, saving €7339 per life year gained, as compared to €3330 when only identified carriers are treated. In sensitivity analysis, outcomes are susceptible to the sensitivity of the screening test and the efficacy of treatment. Reductions in these parameters would reduce the cost-effectiveness of scenarios in which treatment is based on screening. When only identified S. aureus carriers are treated costs of screening should be less than €6.23 to become the dominant strategy.Conclusions
Preoperative screening and eradication of S. aureus carriage to prevent deep-seated S. aureus infections saves both life years and medical costs at the same time, although treating all patients without screening is the dominant strategy, resulting in most health gains and largest savings. 相似文献10.
Alten R Gram H Joosten LA van den Berg WB Sieper J Wassenberg S Burmester G van Riel P Diaz-Lorente M Bruin GJ Woodworth TG Rordorf C Batard Y Wright AM Jung T 《Arthritis research & therapy》2008,10(3):R67