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151.
Active matrix metalloproteinases and degraded collagen are observed in disease states, such as atherosclerosis. To examine whether degraded collagen fragments have distinct effects on vascular smooth muscle cells (SMC), collagenase-digested type I collagen was added to cultured human arterial SMC. After addition of collagen fragments, adherent SMC lose their focal adhesion structures and round up. Analysis of components of the focal adhesion complex demonstrates rapid cleavage of the focal adhesion kinase (pp125(FAK)), paxillin, and talin. Cleavage is suppressed by inhibitors of the proteolytic enzyme, calpain I. In vitro translated pp125(FAK) is a substrate for both calpain I- and II-mediated processing. Mapping of the proteolytic cleavage fragments of pp125(FAK) predicts a dissociation of the focal adhesion targeting (FAT) sequence and second proline-rich domain from the tyrosine kinase domain and integrin-binding sequence. Coimmunoprecipitation studies confirm that the ability of pp125(FAK) to associate with paxillin, vinculin, and p130cas is significantly reduced in SMC treated with degraded collagen fragments. Further, there is a significant reduction in the association of intact pp125(FAK) with the cytoskeletal fraction, while pp125(FAK) cleavage fragments appear in the cytoplasm in SMC treated with degraded collagen fragments. Integrin-blocking studies indicate that integrin-mediated signals are involved in degraded collagen induction of pp125(FAK) cleavage. Thus, collagen fragments induce distinct integrin signals that lead to initiation of calpain-mediated cleavage of pp125(FAK), paxillin, and talin and dissolution of the focal adhesion complex. 相似文献
152.
Trichinella spiralis: behavior, structure, and biochemistry of larvae following exposure to components of the host enteric environment 总被引:1,自引:0,他引:1
Four layers are present on the surface of infective larvae of Trichinella spiralis isolated from host muscle in pepsin-HCl. Trypsin treatment of pepsin-HCl isolated worms caused partial degradation and removal of large patches of the two outer surface layers. Following exposure to bile, only traces of the outer layers remained on the worms surface. These changes in the worm surface were accompanied by a shift from Type I behavior, typical of pepsin-HCl isolated larvae, to Type II behavior, (snakelike) following exposure to either trypsin or bile. Worm behavior was also temperature dependent. Type I behavior was typical of worms maintained at room temperature regardless of treatment, while Type II behavior displayed by worms held at 37 C was treatment dependent. The absorption of in vitro glucose or beta-methyl-D-glucoside was lowest in pepsin-HCl isolated first stage infective larvae, significantly higher in trypsin treated worms and greatest in worms following exposure to bile. Sugar uptake by worms isolated from the host small intestine after 1 hr of enteral infection was similar to that seen in worms isolated from host muscle in pepsin-HCl. Sugar uptake in vitro in worms 2 hr following enteral infection was similar to worms following exposure to bile. The highest levels of sugar absorption in vitro occurred in worms which had resided in the small intestine for 3 hr. The lowest rates of incorporation of label into worm tissues was seen in 1 hr enteral and pepsin-HCl isolated worms. Infective larvae treated with trypsin or bile incorporated significantly greater amounts of label than the two former groups. The highest levels of incorporation of label into worm tissues was seen in 3 hr enteral worms. These findings support the view that trypsin, bile, and temperature serve as environmental cues which lead to alteration of the parasite's behavioral and nutritional status. 相似文献
153.
Raines RT 《Protein science : a publication of the Protein Society》2006,15(5):1219-1225
Collagen is the most abundant protein in animals. The conformational stability of the collagen triple helix is enhanced by the hydroxyl group of its prevalent (2S,4R)-4-hydroxyproline residues. For 25 years, the prevailing paradigm had been that this enhanced stability is due to hydrogen bonds mediated by bridging water molecules. We tested this hypothesis with synthetic collagen triple helices containing 4-fluoroproline residues. The results have unveiled a wealth of stereoelectronic effects that contribute markedly to the stability of collagen, as well as other proteins. This new understanding is leading to synthetic collagens for a variety of applications in biotechnology and biomedicine. 相似文献
154.
Understanding how photosynthetic capacity acclimatises when plants are grown in an atmosphere of rising CO2 concentrations will be vital to the development of mechanistic models of the response of plant productivity to global environmental change. A limitation to the study of acclimatisation is the small amount of material that may be destructively harvested from long-term studies of the effects of elevation of CO2 concentration. Technological developments in the measurement of gas exchange, fluorescence and absorption spectroscopy, coupled with theoretical developments in the interpretation of measured values now allow detailed analyses of limitations to photosynthesisin vivo. The use of leaf chambers with Ulbricht integrating spheres allows separation of change in the maximum efficiency of energy transduction in the assimilation of CO2 from changes in tissue absorptance. Analysis of the response of CO2 assimilation to intercellular CO2 concentration allows quantitative determination of the limitation imposed by stomata, carboxylation efficiency, and the rate of regeneration of ribulose 1:5 bisphosphate. Chlorophyll fluorescence provides a rapid method for detecting photoinhibition in heterogeneously illuminated leaves within canopies in the field. Modulated fluorescence and absorption spectroscopy allow parallel measurements of the efficiency of light utilisation in electron transport through photosystems I and IIin situ.Abbreviations A
net rate of CO2 uptke per unit leaf area (µmol m–2 s–1)
- Asat
light-saturated A
- A820
change in absorptance of PSI on removal of illumination (OD)
- c
CO2 concentration in air (µmol mol–1)
- ca
c in the bulk air; ci, c in the intercellular spaces
- ce
carboxylation efficiency (mol m–2 s–1)
- E
transpiration per unit leaf area (mol m–2 s–1)
- F
fluorescence emission of PSII (relative units)
- Fm
maximal level of F
- Fo
minimal level of F upon illumination when PSII is maximally oxidised
- Fs
the steady-state F following the m peak
- Fv
the difference between Fm and Fo
- F'm
maximal F' generated after the m peak by addition of a saturating light pulse
- F'o
the minimal level of F' after the m peak determined by re-oxidising PSII by far-red light
- g1
leaf conductance to CO2 diffusion in the gas phase (mol m–2 s–1)
- g'1
leaf conductance to water vapour diffusion in the gas phase (mol m–2 s–1)
- kc and ko
the Michaelis constants for CO2 and O2, respectively, (µmol mol–1);
- Jmax
the maximum rate of regeneration of rubP (µmol m–2 s–1)
- l
stomatal limitation to CO2 uptake (dimensionless, 0–1)
- LCP
light compensation point of photosynthesis (µmol m–2 s–1)
- oi
the intercellular O2 concentration (mmol mol–1)
- Pi
cytosol inorganic phosphate concentration
- PSI
photosystem I
- PSII
photosystem II
- Q
photon flux (µmol m–2 s–1)
- Qabs
Q absorbed by the leaf
- rubisCO
ribulose 1:5 bisphosphate carboxylase/oxygenase; rubP, ribulose 1:5 bisphosphate; s, projected surface area of a leaf (m2)
- Vc,max
is the maximum rate of carboxylation (µmol m–2 s–1)
- Wc
the rubisCO limited rate of carboxylation (µmol m–2 s1)
- Wj
the electron transport limited rate of regeneration of rubP (µmol m–2 s–1)
- Wp
the inorganic phosphate limited rate of regeneration of rubP (µmol m–2 s–1)
-
absorptance of light (dimensionless, 0–1)
- a
of standard black absorber 1, of leaf
- s
of integrating sphere walls
- , CO2
compensation point of photosynthesis (µmol mol–1)
-
the specificity factor for rubisCO carboxylation (dimensionless)
- ,
convexity of the response of A to Q (dimensionless 0–1)
-
the quantum yield of photosynthesis on an absorbed light basis (A/Qabs; dimensionless)
-
the quantum yield of photosynthesis on an incident light basis (A/Q; dimensionless)
- app
the maximum
- m
the maximum
- m,app
the photochemical efficiency of PSII (dimensionless, 0–1)
- PSII,m
the maximum 相似文献
155.
Platelet-derived growth factor (PDGF) is secreted by several cells that participate in the process of atherogenesis, including arterial wall monocyte-derived macrophages. Macrophages in human and non-human primate lesions have recently been demonstrated to contain PDGF-B chain protein in situ. In developing lesions of atherosclerosis, macrophages take up and metabolize modified lipoproteins, leading to lipid accumulation and foam cell formation. Oxidatively modified low density lipoproteins (LDL) have been implicated in atherogenesis and have been demonstrated in atherosclerotic lesions. The effects of the uptake of various forms of modified LDL on PDGF gene expression, synthesis, and secretion in adherent cultures of human blood monocyte-derived macrophages were examined. LDL oxidized in a cell-free system in the presence of air and copper inhibited the constitutive expression of PDGF-B mRNA and secretion of PDGF in a dose-dependent fashion. Oxidatively modified LDL also attenuated lipopolysaccharide-induced PDGF-B mRNA expression. These changes were unrelated to the mechanism of lipid uptake and the degree of lipid loading and were detectable within 2 h of exposure to oxidized LDL. The degree of inhibition of both basal and lipopolysaccharide-induced PDGF-B-chain expression increased with the extent of LDL oxidation. Monocyte-derived macrophages exposed to acetylated LDL or LDL aggregates accumulated more cholesterol than cells treated with oxidized LDL, but PDGF expression was not consistently altered. Thus, uptake of a product or products of LDL oxidation modulates the expression and secretion of one of the principal macrophage-derived growth factors, PDGF. This modulation may influence chemotaxis and mitogenesis of smooth muscle cells locally in the artery wall during atherogenesis. 相似文献
156.
Shiaoman Chao Christine A. Raines Marian Longstaff Peter J. Sharp Michael D. Gale Tristan A. Dyer 《Molecular & general genetics : MGG》1989,218(3):423-430
Summary Homologous probes for the wheat coding sequences of the enzymes phosphoribulokinase, phosphoglycerate kinase (both chloroplast and cytosolic forms), chloroplast fructose-1,6-bisphosphatase and the small subunit of ribulose-1,5-bisphosphate carboxylase were used to determine the copy number and chromosomal location of the genes encoding these enzymes by restriction fragment length polymorphism analysis. Heterologous probes were similarly used to characterize the genes for the enzymes glyceraldehyde phosphate dehydrogenase (both chloroplast and cytosolic forms), phosphoenolpyruvate carboxylase and pyruvate, orthophosphate dikinase. Several of the genes are present in single copies per haploid genome, and the different enzymes are encoded by loci dispersed on different chromosomes. The significance of these findings is discussed in relation to gene expression and control of copy number. 相似文献
157.
Graham J. C. Underwood Matthew Boulcott Christine A. Raines Keith Waldron 《Journal of phycology》2004,40(2):293-304
Marine benthic diatoms excrete large quantities of extracellular polymeric substances (EPS), both as a function of their motility system and as a response to environmental conditions. Diatom EPS consists predominantly of carbohydrate‐rich polymers and is important in the ecology of cells living on marine sediments. Production rates, production pathways, and monosaccharide composition of water‐soluble (colloidal) carbohydrates, EPS, and intracellular storage carbohydrate (glucans) were investigated in the epipelic (mud‐inhabiting) diatoms Cylindrotheca closterium (Ehrenburg), Navicula perminta (Grün.) in Van Heurck, and Amphora exigua Greg. under a range of experimental conditions simulating aspects of the natural environment. Cellular rates of colloidal carbohydrate, EPS, and glucan production were significantly higher during nutrient‐replete compared with nutrient‐limited growth for all three species. The proportion of EPS in the extracellular carbohydrate pool increased significantly (to 44%–69%) as cells became nutrient limited. Cylindrotheca closterium produced two types of EPS differing in sugar composition and production patterns. Nutrient‐replete cells produced a complex EPS containing rhamnose, fucose, xylose, mannose, galactose, glucose, and uronic acids. Nutrient‐limited cells produced an additional EPS containing mannose, galactose, glucose, and uronic acids. Both EPS types were produced under illuminated and darkened conditions. 14C‐labeling revealed immediate production of 14C‐glucan and significant increases in 14C‐EPS between 3 and 4 h after addition of label. The glucan synthesis inhibitor 2,6‐dichlorobenzonitrile significantly reduced 14C‐colloidal carbohydrate and 14C‐EPS. The glucanase inhibitor P‐nitrophenyl β‐d ‐glucopyranoside resulted in accumulation of glucan within cells and lowered rates of 14C‐colloidal and 14C‐EPS production. Cycloheximide prevented glucan catabolism, but glucan production and EPS synthesis were unaffected. 相似文献
158.
Purification of PDGF-AB and PDGF-BB from human platelet extracts and identification of all three PDGF dimers in human platelets 总被引:21,自引:0,他引:21
We have developed a panel of monoclonal antibodies to platelet-derived growth factor (PDGF) which have variable specificities for the three dimeric forms of the molecule (AA, AB, and BB). We have used these antibodies to detect and immunoaffinity purify the individual dimers from human platelet rich plasma. Extracts of outdated platelet preparations were initially chromatographed over CM-Sepharose and then passed over the Sepharose-coupled monoclonal antibodies in series in selectively isolate the three dimeric forms of PDGF. The PDGF eluted from the affinity columns was subsequently further purified by reversed-phase HPLC. From 300 units of outdated platelet preparations, we purified 58 micrograms of PDGF-BB and 140 micrograms of PDGF-AB. Using the monoclonal antibodies to develop PDGF dimer-specific ELISAs, it was observed that all three PDGF dimer forms are present in fresh human platelet extracts and that the ratios of the three dimer forms vary depending upon the extraction conditions used. The identification of all three PDGF dimer forms in human platelets point to the need to view PDGF isolated from human platelets by conventional techniques as a mixture of all three forms and not solely as PDGF-AB. 相似文献
159.
Christine A. Raines Philip R. Horsnell Caroline Holder Julie C. Lloyd 《Plant molecular biology》1992,20(6):1143-1148
A full-length cDNA clone encoding carbonic anhydrase was isolated from an Arabidopsis thaliana (Columbia) leaf library. Comparison of the derived amino acid sequence obtained from this clone with those of pea and spinach reveals a considerable degree of identity. The carbonic anhydrase cDNA was used to probe the level of RNA encoding this protein in the leaves of plants grown in elevated CO2 (660 ppm). We have found that under these conditions the steady-state level of carbonic anhydrase mRNA was increased in comparison with control plants grown in normal atmospheric concentrations of CO2 (330 ppm). This raises the intruiging possibility that there exists in higher plants a mechanism for perceiving and responding to changes in environmental CO2 concentrations at the genetic level. 相似文献
160.