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221.
In fitting atomic structures into EM maps, it often happens that the map corresponds to a different conformation of the structure. We have developed a new methodology to handle these situations that preserves the covalent geometry of the structure and allows the modeling of large deformations. The first goal is achieved by working in generalized coordinates (positional and internal coordinates), and the second by avoiding harmonic potentials. Instead, we use dampers (shock absorbers) between every pair of atoms, combined with a force field that attracts the atomic structure toward incompletely occupied regions of the EM map. The trajectory obtained by integrating the resulting equations of motion converges to a conformation that, in our validation cases, was very close to the target atomic structure. Compared to current methods, our approach is more efficient and robust against wrong solutions and to overfitting, and does not require user intervention or subjective decisions. Applications to the computation of transition pathways between known conformers, homology and loop modeling, as well as protein docking, are also discussed. 相似文献
222.
Aucamp JP Martinez-Torres RJ Hibbert EG Dalby PA 《Biotechnology and bioengineering》2008,99(6):1303-1310
We have previously developed a rapid microplate-based approach for measuring the denaturation curves by intrinsic tryptophan fluorescence for simple monomeric and two-state unfolding proteins. Here we demonstrate that it can accurately resolve the multiple conformational transitions that occur during the denaturation of a complex multimeric and cofactor associated protein. We have also analyzed the effect of two active-site mutations, D381A and Y440A upon the denaturation pathway of transketolase using intrinsic fluorescence measurements, and we compare the results from classical and microplate-based instrumentation. This work shows that the rapid assay is able to identify changes in the denaturation pathway, due to mutations or removal of cofactors, which affect the stability of the native and intermediate states. This would be of significant benefit for the directed evolution of protein stability, optimizing enzyme stability under biocatalytic process conditions, and also for engineering specific transitions in protein unfolding pathways. 相似文献
223.
State of the art docking algorithms predict an incorrect binding pose for about 50-70% of all ligands when only a single fixed receptor conformation is considered. In many more cases, lack of receptor flexibility results in meaningless ligand binding scores, even when the correct pose is obtained. Incorporating conformational rearrangements of the receptor binding pocket into predictions of both ligand binding pose and binding score is crucial for improving structure-based drug design and virtual ligand screening methodologies. However, direct modeling of protein binding site flexibility remains challenging because of the large conformational space that must be sampled, and difficulties remain in constructing a suitably accurate energy function. Here we show that using multiple fixed receptor conformations, either experimentally determined by crystallography or NMR, or computationally generated, is a practical shortcut that may improve docking calculations. In several cases, such an approach has led to experimentally validated predictions. 相似文献
224.
Algal contact as a trigger for coral disease 总被引:4,自引:0,他引:4
Maggy M. Nugues Garriet W. Smith Ruben J. van Hooidonk Maria I. Seabra Rolf P. M. Bak 《Ecology letters》2004,7(10):919-923
Diseases are causing alarming declines in reef‐building coral species, the foundation blocks of coral reefs. The emergence of these diseases has occurred simultaneously with large increases in the abundance of benthic macroalgae. Here, we show that physical contact with the macroalga Halimeda opuntia can trigger a virulent disease known as white plague type II that has caused widespread mortality in most Caribbean coral species. Colonies of the dominant coral Montastraea faveolata exposed to algal transplants developed the disease whereas unexposed colonies did not. The bacterium Aurantimonas coralicida, causative agent of the disease, was present on H. opuntia sampled close to, and away from diseased corals, indicating that the alga serves as a reservoir for this pathogen. Our results suggest that the spread of macroalgae on coral reefs could account for the elevated incidence of coral diseases over past decades and that reduction of macroalgal abundance could help control coral epizootics. 相似文献
225.
Deborah B Kaufman Marc E Hentsch George A Baumbach Joseph A Buettner Christopher A Dadd Ping Y Huang David J Hammond Ruben G Carbonell 《Biotechnology and bioengineering》2002,77(3):278-289
An affinity resin containing the peptide ligand Phe-Leu-Leu-Val-Pro-Leu (FLLVPL) has been developed for the purification of fibrinogen. The ligand was identified by screening a solid-phase combinatorial peptide library using an immunostaining technique. The specific binding of fibrinogen to the ligand has been characterized by isothermal calorimetry and adsorption isotherms and is dominated by both hydrophobic interactions and ionic interactions with the N-terminal free amino group. The effective association constant of fibrinogen was substantially higher when the peptide was immobilized on the resin than in solution; moreover, it increased with increasing peptide density, suggesting a cooperative binding effect. A low ionic strength buffer at pH 4 was used successfully to elute adsorbed fibrinogen from the column with high purity, retention of factor XIII crosslinking activity, and minimal, if any, loss of biological function. This general approach to ligand selection and characterization can be used to develop peptide ligands for the affinity purification of diverse proteins on a large scale. 相似文献
226.
Goodness of fit of biplots and correspondence analysis 总被引:3,自引:0,他引:3
227.
Riad Efendiev Alejandro M Bertorello Ruben Zandomeni Angel R Cinelli Carlos H Pedemonte 《The Journal of biological chemistry》2002,277(13):11489-11496
We tested the hypothesis that the level of intracellular sodium modulates the hormonal regulation of the Na(+),K(+)-ATPase activity in proximal tubule cells. By using digital imaging fluorescence microscopy of a sodium-sensitive dye, we determined that the sodium ionophore monensin induced a dose-specific increase of intracellular sodium. A correspondence between the elevation of intracellular sodium and the level of dopamine-induced inhibition of Na(+),K(+)-ATPase activity was determined. At basal intracellular sodium concentration, stimulation of cellular protein kinase C by phorbol 12-myristate 13-acetate (PMA) promoted a significant increase in Na(+),K(+)-ATPase activity; however, this activation was gradually reduced as the concentration of intracellular sodium was increased to become a significant inhibition at concentrations of intracellular sodium higher than 16 mm. Under these conditions, PMA and dopamine share the same signaling pathway to inhibit the Na(+),K(+)-ATPase. The effects of PMA and dopamine on the Na(+),K(+)-ATPase activity and the modulation of these effects by different intracellular sodium concentrations were not modified when extracellular and intracellular calcium were almost eliminated. These results suggest that the level of intracellular sodium modulates whether hormones stimulate, inhibit, or have no effect on the Na(+),K(+)-ATPase activity leading to a tight control of sodium reabsorption. 相似文献
228.
A field experiment was carried out in October 1998 during active upwelling in a coastal area off the Mejillones Peninsula (23° S). Zooplankton was sampled at day and night, during two subsequent days at 4 stations inside and outside of the upwelling plume. Three depth strata were sampled: 0–20 m, 20–80 m and 80–200 m. Oceanographic data were obtained in a grid of 23 stations using a CTDO, a fluorometer and a Doppler current meter. Calanus chilensis was mostly represented by late stages, i.e. copepodid C5 and adult males and females. There were no day/night effects on vertical distribution, and abundance was significantly higher inside the upwelling plume in the upper 20-m layer at nearly 14 ind. m–3, compared to ca. 5 ind. m–3 outside the upwelling plume. Temperature at 10 m depth and biomass, estimated from stage numbers and their mean dry weights, were used to estimate growth and daily production of Calanus at temperature-dependent rates. The potential loss of biomass from the upwelling center because of advection in the upwelling plume was estimated from current data in the Ekman layer and biomass density. The mean cross-shelf component of the current was estimated at 10.4 km d–1 within the upwelling plume. This yielded a loss of biomass of 9.7 mg dry weight m–2 . Production, estimated by a temperature-dependent approach, ranged between 44 and 35 mg dry weight m–2 d–1, at mean temperatures of 14.6 °C and 15.8 °C inside and outside of the upwelling plume respectively. Within the plume, as much as 22% of daily production may be advected offshore. However, a higher concentration of biomass in the upwelling plume allowed a greater production compared to surrounding areas. A mass balance approach suggests that advective losses may not have a major impact on the C. chilensis population, because of very high daily production at temperature-dependent rates. 相似文献
229.
230.
Gianfranco Sebastio Ornella Castiglione Barbara Incerti Donatello Salvatore Francesca Santamaria 《Human genetics》1990,85(4):430-431
Summary Fifty one independent cystic fibrosis (CF) families originating from a restricted area of Southern Italy (Campania) have been
analyzed for KM19 and XV2c haplotypes and the ΔF508 mutation: 54% of the total CF chromosomes show the ΔF508 mutation. No
significative correlations were obtained when clinical score, radiological score,Pseudomonas colonization, or clinical symptoms at presentation were matched with the presence or absence of the ΔF508 mutation. 相似文献