全文获取类型
收费全文 | 892篇 |
免费 | 77篇 |
国内免费 | 3篇 |
专业分类
972篇 |
出版年
2021年 | 17篇 |
2020年 | 7篇 |
2019年 | 11篇 |
2018年 | 18篇 |
2017年 | 13篇 |
2016年 | 25篇 |
2015年 | 38篇 |
2014年 | 28篇 |
2013年 | 74篇 |
2012年 | 39篇 |
2011年 | 47篇 |
2010年 | 30篇 |
2009年 | 30篇 |
2008年 | 44篇 |
2007年 | 38篇 |
2006年 | 41篇 |
2005年 | 30篇 |
2004年 | 26篇 |
2003年 | 27篇 |
2002年 | 19篇 |
2001年 | 27篇 |
2000年 | 23篇 |
1999年 | 27篇 |
1998年 | 18篇 |
1997年 | 18篇 |
1996年 | 11篇 |
1995年 | 7篇 |
1994年 | 6篇 |
1993年 | 6篇 |
1992年 | 10篇 |
1991年 | 16篇 |
1990年 | 15篇 |
1989年 | 5篇 |
1988年 | 13篇 |
1987年 | 12篇 |
1986年 | 13篇 |
1985年 | 18篇 |
1984年 | 8篇 |
1983年 | 8篇 |
1982年 | 7篇 |
1981年 | 8篇 |
1980年 | 4篇 |
1979年 | 11篇 |
1976年 | 6篇 |
1974年 | 4篇 |
1973年 | 5篇 |
1972年 | 5篇 |
1971年 | 4篇 |
1969年 | 6篇 |
1967年 | 5篇 |
排序方式: 共有972条查询结果,搜索用时 15 毫秒
1.
B M Foxwell D C Blakey A N Brown T A Donovan P E Thorpe 《Biochimica et biophysica acta》1987,923(1):59-65
Deglycosylation of ricin may be necessary to prevent the entrapment of antibody-ricin conjugates in vivo by cells of the reticuloendothelial system which have receptors that recognise the oligosaccharide side chains on the A- and B-chains of the toxin. Carbohydrate-deficient ricin was therefore prepared by recombining the A-chain, which had been treated with alpha-mannosidase, with the B-chain, which had been treated with endoglycosidase H or alpha-mannosidase or both. By recombining treated and untreated chains, a series of ricin preparations was made having different carbohydrate moieties. The removal of carbohydrate from the B-chain did not affect the ability of the toxin to agglutinate erythrocytes, and alpha-mannosidase treatment of the A-chain did not affect its ability to inactivate ribosomes. The toxicity of ricin to cells in culture was only reduced in those preparations containing B-chain that had been treated with alpha-mannosidase, when a 75% decrease in toxicity was observed. The toxicity of the combined ricin preparation to mice varied from double to half that of native ricin, depending on the chain(s) treated and the enzymes used. Removal of carbohydrate greatly reduced the hepatic clearance of the toxin and the levels of toxin in the blood were correspondingly higher. These results suggest that antibody-ricin conjugates prepared from deglycosylated ricin would be cleared more slowly by the liver, inflict less liver damage, and have greater opportunity to reach their target. 相似文献
2.
The interaction of Ca2+ with the calmodulin-sensitive adenylate cyclase from Bordetella pertussis 总被引:2,自引:0,他引:2
H R Masure D J Oldenburg M G Donovan R L Shattuck D R Storm 《The Journal of biological chemistry》1988,263(14):6933-6940
Bordetella pertussis, the etiologic agent of whooping cough, produces a calmodulin-sensitive adenylate cyclase which elevates intracellular cAMP in a variety of eucaryotic cells. Exogenous calmodulin added to the partially purified adenylate cyclase has been shown to inhibit invasion of animal cells by this enzyme (Shattuck, R. L., and Storm, D. R. (1985) Biochemistry 24, 6323-6328). In this study, several properties of the calmodulin-sensitive adenylate cyclase are shown to be influenced by Ca2+ in the absence of calmodulin. The presence or absence of Ca2+ during QAE-Sephadex ion exchange chromatography produced two distinct chromatographic patterns of adenylate cyclase activity. Two different forms of the enzyme (Pk1 and Pk2EGTA) were isolated by this procedure. Pk1 adenylate cyclase readily elevated intracellular cAMP levels in mouse neuroblastoma cells (N1E-115) while Pk2EGTA adenylate cyclase had no effect on cAMP levels in these cells. Gel exclusion chromatography of Pk1 adenylate cyclase gave apparent Stokes radii (RS) of 43.5 A (+/- 1.3) in the presence of 2 mM CaCl2 and 33.8 A (+/- 0.94) in the presence of 2 mM EGTA [( ethylenebis (oxyethylenenitrilo)]tetraacetic acid). These Stokes radii are consistent with molecular weights of 104,000 (+/- 6,400) and 61,000 (+/- 3,600), respectively. Pk2EGTA adenylate cyclase had an apparent RS of 33.0 (+/- 1.2) (Mr = 60,600 (+/- 2,800] in the presence of Ca2+ or excess EGTA. At 60 degrees C, Pk1 adenylate cyclase exhibited a Ca2+-dependent heat stability with a half-life for loss of enzyme activity of 10.3 min in 5 mM CaCl2 and a half-life of 2.8 min in the presence of 0.1 microM CaCl2. The stability of Pk2EGTA adenylate cyclase was not affected by changes in free Ca2+. The adenylate cyclase preparations described above were submitted to sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis, and enzyme activity was recovered from gel slices by extraction with detergent containing buffers. The catalytic subunit isolated from SDS-polyacrylamide gels was activated 7-fold in the presence of Ca2+ with maximum activity observed at 1 microM free Ca2+. With both preparations, the apparent molecular weight of the catalytic subunit on SDS gels was 51,000 in the presence of 2 mM CaCl2 and 45,000 in the presence of 2 mM EGTA. The catalytic subunit of the enzyme was purified to apparent homogeneity by preparative SDS-polyacrylamide gel electrophoresis and resubmitted to SDS gel electrophoresis in the presence or absence of free Ca2+. The purified catalytic subunit also exhibited a Ca2+-dependent shift in its mobility on SDS gels.(ABSTRACT TRUNCATED AT 400 WORDS) 相似文献
3.
4.
Seasonal carbon isotope discrimination in a grassland community 总被引:11,自引:0,他引:11
Mark P. Smedley Todd E. Dawson Jonathan P. Comstock Lisa A. Donovan Dorothy E. Sherrill Craig S. Cook James R. Ehleringer 《Oecologia》1991,85(3):314-320
Summary Grassland communities of arid western North America are often characterized by a seasonal increase in ambient temperature and evaporative demand and a corresponding decline in soil moisture availability. As the environment changes, particular species could respond differently, which should be reflected in a number of physiological processes. Carbon isotope discrimination varies during photosynthetic activity as a function of both stomatal aperture and the biochemistry of the fixation process, and provides an integrated measure of plant response to seasonal changes in the environment. We measured the seasonal course of carbon isotope discrimination in 42 grassland species to evaluate changes in gas exchange processes in response to these varying environmental factors. The seasonal courses were then used to identify community-wide patterns associated with life form, with phenology and with differences between grasses and forbs. Significant differences were detected in the following comparisons: (1) Carbon isotope discrimination decreased throughout the growing season; (2) perennial species discriminated less than annual species; (3) grasses discriminated less than forbs; and (4) early flowering species discriminated more than the later flowering ones. These comparisons suggested that (1) species active only during the initial, less stressful months of the growing season used water less efficiently, and (2) that physiological responses increasing the ratio of carbon fixed to water lost were common in these grassland species, and were correlated with the increase in evaporative demand and the decrease in soil moisture. 相似文献
5.
Influence of dilution on the physical state of model bile systems: NMR and quasi-elastic light-scattering investigations 总被引:1,自引:0,他引:1
Multinuclear (1H and 31P) nuclear magnetic resonance (NMR) spectroscopy and quasi-elastic light scattering have been used to characterize molecular aggregates formed in dilute sodium taurocholate--egg lecithin solutions. When mixed micelles (1.25 g/dL) are diluted with 150 mM aqueous sodium chloride, light-scattering measurements suggest a transformation from mixed micelles to unilamellar vesicle species. Decreased 1H NMR line widths for bile salt resonances are consistent with predominance of a monomer form. The concurrent appearance of a second phospholipid choline methyl resonance indicates two types of phospholipid environment in slow chemical exchange: this behavior is consistent with small unilamellar vesicles. The appearance of bilayer vesicles in dilute model bile solutions is confirmed by addition of a lanthanide shift reagent (Pr3+), which splits the 1H or 31P head-group peak into two components with distinct chemical shift sensitivities. These mixed micelle and vesicle aggregates are also distinguished by their susceptibility to the lipolytic enzyme phospholipase A2 from cobra venom. 相似文献
6.
A charge coupled device-based image cytophotometry system for quantitative histochemistry and cytochemistry 总被引:1,自引:0,他引:1
A rapid, semiautomated cytophotometry system for quantitative histochemistry and cytochemistry was constructed. The system consists of a Fairchild charge coupled device (CCD) image camera, a Zeiss Universal microscope, a Datacube analog to digital converter, and a digital Equipment Corporation LSI 11/23 computer operating under RT-11. Computer programs were written in FORTRAN and the MACRO assembly language for the acquisition of data from the CCD device. These data were then used for image segmentation, image display, and calculation of total optical density, perimeter, cell area, and several shape features. The reproducibility of measurement made with the CCD-based cytophotometry system was tested by repeated measurements. The coefficient of variation was estimated to be 1.7% for total optical density and 0.9% for cell area. The CCD-based cytophotometry system was further evaluated by comparing results with measurements made on the same cells with a scanning stage cytophotometer using the HIDACSYS computer programs. Correlation coefficients of 0.96 for total optical density and 0.91 for cell area were obtained between the two systems. We conclude that the high-speed, dimensional stability, small size, and linearity of the CCD-based cytophotometry system will make it useful for quantitative histochemistry and cytochemistry. 相似文献
7.
Requirements for the translocation of diphtheria toxin fragment A across lipid membranes 总被引:2,自引:0,他引:2
The translocation of the enzymatic moiety of diphtheria toxin, fragment A, across the membranes of pure lipid vesicles was demonstrated. A new assay, which employed vesicles made to contain radiolabeled NAD and elongation factor-2, was used to measure the appearance of the enzymatic activity of the A fragment in the vesicles. When the vesicles were exposed to a low-pH medium in the presence of diphtheria toxin, small molecules, such as NAD, escaped into the extravesicular medium, whereas large molecules mostly remained inside the vesicles. The vesicle-entrapped elongation factor-2 became ADP-ribosylated, indicating the entry of fragment A into the vesicle. The translocation of the A fragment depended upon the pH of the medium, being negligible at pH greater than 7.0 and maximal at pH 4.5. The entire toxin molecule was needed for function; neither the A fragment nor the B fragment alone was able to translocate itself across and react with the sequestered substrates. After exposure of the toxin to low pH, the entry of the A fragment was rapid, being virtually complete within 2-3 min at pH 5.5, and within 1 min at pH 4.7. Translocation occurred in the absence of any protein in the vesicle membrane. These results are consistent with the notion that the diphtheria toxin molecule enters the cytoplasm of a cell by escaping from an acidic compartment such as an endocytic vesicle. 相似文献
8.
Isolation and characterization of a large,neurite-associated glycoconjugate from neuroblastoma cells
下载免费PDF全文
![点击此处可从《The Journal of cell biology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
A high molecular weight glycoconjugate has been isolated from neurite-producing neuronal tumor cells in culture and has been designated as I(0) based on its elution characteristics in gel filtration chromatography. This molecule cannot be found in a variety of nonneuronal cells. I(0) is found in the substratum-attached material or cell fraction of neurite-producing neuroblastoma cells, depending upon culture conditions. It is found in the substratum-bound fraction of B104 rat neuroblastoma cells during serum starvation and in the EGTA-detached cell fraction of B104 cells grown in chemically defined N2 medium. It occurs only in the cell fraction of the human neuroblastoma line Platt. Examination of behavioral variants of the B104 rat line further strengthens the association of I(0) with neurite production; the constitutive neurite-producing E(R)B9 variant contains I(0) while the non-neurite-producing E(R)A11 variant does not. I(0) is large, eluting in the void volume of sepharose-CL2B columns. Radioiodination of intact cells with lactoperoxidase shows I(0) to be a cell surface component. Metabolic radiolabeling studies show that it contains a high proportion of polysaccharide to protein, does not contain mannose, and is unsulfated. Alkaline borohydride reduction release two size classes of large polysaccharide chain. The alkaline reduction results, along with the mannose incorporation studies, show the presence of O-glycosidic linkages and few, if any, N-linkages. Resistance to nitrous acid deamination, insensitivity to glycosaminoglycan lyases, and the absence of sulfation, indicate that I(0) does not contain the glycosaminoglycans hyaluronic acid, chondroitin-, dermatan-, or heparin- sulfates. Affinity column chromatography reveals high binding affinity of I(0) to polyornithine and no binding to gelatin (collagen) or the glycosaminoglycans hyaluronate and heparin. These studies describe a unique high molecular weight glycoconjugate on the surface of neurite-producing neuroblastoma cell lines from two species. 相似文献
9.
Summary Uranyl sulphate (0.2–0.9 mM) inhibited ferrous iron oxidation by growing cultures ofThiobacillus ferrooxidans. The addition of 5–100 mM uranium to the cultures caused immediate cessation of carbon dioxide fixation, rapid loss of viability
and gradual depression of ferrous iron oxidation. Virtually no uranium was found in washed cells grown in the presence of
subtoxic to toxic amounts of uranyl sulphate. Uranium-poisoned organisms appeared plasmolyzed in electron micrographs. Cultures
tolerant to 5 mM UO2
2+ were develoepd by successive subculturing in increased uranium concentrations. The tolerance was maintained during subculturing
in uranium-free medium. Frequency of mutants resistant to 1.0 and 1.5 mM UO2
2+ was 1 per 1.3×106 and 1 per 9.0×108, respectively. The frequency was increased in the presence of 15–150 mM nickel, zinc and manganese. In liquid cultures, bivalent
cations and EDTA alleviated the toxicity of 2 mM uranyl sulphate. 相似文献
10.
Lisa A. Donovan James H. Richards Matthew W. Muller 《American journal of botany》1996,83(12):1637-1646
At Mono Lake, California, we investigated field water relations, leaf and xylem chemistry, and gas exchange for two shrub species that commonly co-occur on marginally saline soils, and have similar life histories and rooting patterns. Both species had highest root length densities close to the surface and have large tap roots that probably reach ground water at 3.4-5.0 m on the study site. The species differed greatly in leaf water relations and leaf chemistry. Sarcobatus vermiculatus had a seasonal minimum predawn xylem pressure potential (ψpd) of -2.7 MPa and a midday potential (ψmd) of -4.1 MPa. These were significantly lower than for Chrysothamnus nauseosus, which had a minimum ψpd of -1.0 MPa and ψmd of -2.2 MPa. Sarcobatus had leaf Na of up to 9.1 % and K up to 2.7 % of dry mass, and these were significantly higher than for Chrysothamnus which had seasonal maxima of 0.4% leaf Na and 2.4 % leaf K. The molar ratios of leaf K/Na, Ca/Na, and Mg/Na were substantially lower for Sarcobatus than for Chrysothamnus. Xylem ionic contents indicated that both species excluded some Na at the root, but that Chrysothamnus was excluding much more than Sarcobatus. The higher Na content of Sarcobatus leaves was associated with greater leaf succulence, lower calculated osmotic potential, and lower xylem pressure potentials. Despite large differences in water relations and leaf chemistry, these species maintained similar diurnal patterns and rates of photosynthesis and stomatal conductance to water vapor diffusion. Sarcobatus ψpd may not reflect soil moisture availability due to root osmotic and hydraulic properties. 相似文献