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31.
Crystal structure of cholera toxin B-pentamer bound to receptor GM1 pentasaccharide. 总被引:25,自引:5,他引:20 下载免费PDF全文
E. A. Merritt S. Sarfaty F. van den Akker C. L'Hoir J. A. Martial W. G. Hol 《Protein science : a publication of the Protein Society》1994,3(2):166-175
Cholera toxin (CT) is an AB5 hexameric protein responsible for the symptoms produced by Vibrio cholerae infection. In the first step of cell intoxication, the B-pentamer of the toxin binds specifically to the branched pentasaccharide moiety of ganglioside GM1 on the surface of target human intestinal epithelial cells. We present here the crystal structure of the cholera toxin B-pentamer complexed with the GM1 pentasaccharide. Each receptor binding site on the toxin is found to lie primarily within a single B-subunit, with a single solvent-mediated hydrogen bond from residue Gly 33 of an adjacent subunit. The large majority of interactions between the receptor and the toxin involve the 2 terminal sugars of GM1, galactose and sialic acid, with a smaller contribution from the N-acetyl galactosamine residue. The binding of GM1 to cholera toxin thus resembles a 2-fingered grip: the Gal(beta 1-3)GalNAc moiety representing the "forefinger" and the sialic acid representing the "thumb." The residues forming the binding site are conserved between cholera toxin and the homologous heat-labile enterotoxin from Escherichia coli, with the sole exception of His 13. Some reported differences in the binding affinity of the 2 toxins for gangliosides other than GM1 may be rationalized by sequence differences at this residue. The CTB5:GM1 pentasaccharide complex described here provides a detailed view of a protein:ganglioside specific binding interaction, and as such is of interest not only for understanding cholera pathogenesis and for the design of drugs and development of vaccines but also for modeling other protein:ganglioside interactions such as those involved in GM1-mediated signal transduction. 相似文献
32.
33.
The present study describes the biochemical characteristics of an acid -fructosidase (EC 3.2.1.26) purified from the fruit of sweet pepper (Capsicum annuum L.). The soluble form, which constitutes more than 95% of the total activity at pH 4.5, hydrolyzes sucrose, raffinose, and stachyose. Its pH and temperature optima are 4.5 and 55 °C, respectively. Metal cations such as Ag+ and Hg2+ strongly inhibit its activity, suggesting the presence of at least one sulfhydryl group at the catalytic site. After purification of the enzyme by means of ammonium sulfate fractionation, gel chromatography (diethyl-aminoethyl-Sephacel, hydroxylapatite, concanavalin A-Sepharose), and preparative gel electrophoresis, the purified enzyme was shown to be a 42 kDa glycoprotein interacting specifically with concanavalin A. After complete chemical deglycosylation with trifluoromethanesulfonic acid, the molecular weight of the constitutive polypeptide was estimated to be 39 kDa. The enzyme glycans were characterized using both affino- and immunodetection. The enzyme has at least two N-linked oligosaccharide sidechains, one of the high-mannose type, and the other of the complex type. The high-mannose glycan has a low molecular weight (1 kDa), and is responsible for the interaction between the enzyme and concanavalin A. The complex-type glycan has an estimated molecular weight of 2 kDa. It contains one 1 2-linked xylose residue, probably one fucose residue 1 3-linked to the chitobiose unit, and no terminal galactose residue. The two glycans, associated to the 39 kDa polypeptide, constitute the acid -fructosidase of the sweet-pepper fruit.Abbreviations F
-fructosidase
- ConA
concanavalin A
- DEAE
diethylaminoethyl
- DTNB
dithionitrobenzoic acid
- endo F
endo--N-acetylglucosamidase F
- endo H
endo--N-acetylglucosamidase H
- NEM
N-ethylmaleimide
- PCMB
parachloromercurobenzoate
- PNGase
glycopeptide-N-glycosidase
- TFMS
trifluoromethane sulfonic acid
This work was partly supported by a grant from the Commission Permanente de Coopération Franco-Québécoise to L. Faye, and S. Yelle. D. Michaud was a recipient of a graduate scholarship from the Natural Science and Engineering Research Council of Canada. 相似文献
34.
Different CMS sources found in Beta vulgaris ssp maritima: mitochondrial variability in wild populations revealed by a rapid screening procedure 总被引:1,自引:0,他引:1
P. Saumitou-Laprade G. J. A. Rouwendal J. Cuguen F. A. Krens G. Michaelis 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1993,85(5):529-535
Summary Mitochondrial DNA (mtDNA) variation in natural Beta maritima populations has been characterized by way of Southern blot hybridizations of total DNA using non-radioactive probes and chemiluminescent detection. It was found that the previously described N (normal) mitochondrial type could be subdivided into three subtypes. A new mitochondrial genotype (type R) was distinguished in addition to the previously described type S. Both are male-sterile cytoplasms and can produce a. segregation of sexual phenotypes in their progenies depending on the nuclear background. The populations contained at least two to four different mitochondrial genotypes. 相似文献
35.
S. G. Kilian B. A. Prior J. C. du Preez 《World journal of microbiology & biotechnology》1993,9(3):357-360
Low-affinity (K
m=67.6±3.2 mM) and high-affinity (K
m=1.9±1.2 mM) D-xylose transport occur in Candida utilis grown, respectively, on D-glucose or D-xylose. Starvation of glucose-grown cells decreases the K
m value (10.5±2.6 mm). The high-affinity system appearing during starvation required protein synthesis and it was inactivated when cells were exposed to glucose, by a process independent of protein synthesis. High-affinity transport was accompanied by transient alkalinization of yeast suspensions, indicating that it is a proton symport, whereas low-affinity transport was not. Both systems, however, were inhibited by metabolic inhibitors and by replacing H2O in the transport assay with D2O, indicating that both may be proton symports. Glucose and acetic acid also inhibited both high-and low-affinity xylose transport.S.G. Kilian, B.A. Prior and J.C. du Preez are with the Department of Microbiology and Biochemistry, University of the Orange Free State, P.O. Box 339, Bloemfontein 9300, Republic of South Africa 相似文献
36.
Jonathan Reizer Antonio H. Romano Josef Deutscher 《Journal of cellular biochemistry》1993,51(1):19-24
HPr of the Gram-positive bacterial phosphotransferase system (PTS) can be phosphorylated by an ATP-dependent protein kinase on a serine residue or by PEP-dependent Enzyme I on a histidyl residue. Both phosphorylation events appear to influence the metabolism of non-PTS carbon sources. Catabolite repression of the gluconate (gnt) operon of B. subtilis appears to be regulated by the former phosphorylation event, while glycerol kinase appears to be regulated by the latter phosphorylation reaction. The extent of our understanding of these processes will be described. © 1993 Wiley-Liss, Inc. 相似文献
37.
Hiroshi Takeda 《Journal of plant research》1993,106(3):195-200
Matrix sugar composition ofChlorella is species-specifically different. The rigid wall consists of either glucosamine or glucose and mannose. Ruthenium red stainability
and anisotropy of cell wall are either plus or minus species-specifically. The cell wall is specifically degraded by the lytic
enzyme of the cell itself. 相似文献
38.
Viviane Fátima de Oliveira Lucas Figueiredo Gabriel de Oliveira Correia Maria de Fátima Pires da Silva Machado Hugo Zeni Neto Wanderley Dantas dos Santos Claudete Aparecida Mangolin 《Global Change Biology Bioenergy》2023,15(12):1465-1476
The burgeoning cellulosic ethanol industry necessitates advancements in enzymatic saccharification, effective pretreatments for lignin removal, and the cultivation of crops more amenable to saccharification. Studies have demonstrated that natural inhibitors of lignin biosynthesis can enhance the saccharification of lignocellulose, even in tissues generated several months post-treatment. In this study, we applied daidzin (a competitive inhibitor of coniferaldehyde dehydrogenase), piperonylic acid (a quasi-irreversible inhibitor of cinnamate 4-hydroxylase), and methylenedioxy cinnamic acid (a competitive inhibitor of 4-coenzyme A ligase) to 60-day-old crops of two conventional Brazilian sugarcane cultivars and two energy cane clones, bred specifically for enhanced biomass production. The resultant biomasses were evaluated for lignin content and enzymatic saccharification efficiency without additional lignin-removal pretreatments. The treatments amplified the production of fermentable sugars in both the sugarcane cultivars and energy cane clones. The most successful results softened the most recalcitrant lignocellulose to the level of the least recalcitrant of the biomasses tested. Interestingly, the softest material became even more susceptible to saccharification. 相似文献
39.
James A. Bunce 《Physiologia plantarum》1992,86(1):173-179
The short-term stimulation of the net rate of carbon dioxide exchange of leaves by elevated concentrations of CO2 usually observed in C3 plants sometimes does not persist. Experiments were conducted to test whether the patterns of response to the environment during growth were consistent with the hypotheses that photosynthetic adjustment to elevated CO2 concentration is due to (1) feedback inhibition or (2) nutrient stress. Soybean [Glycine max (L.) Merr. cv. Williams] and sugar beet (Best vulgaris L. cv. Mono Hye-4) were grown from seed at 350 and 700 μl? CO2, at 20 and 25°C, at a photon flux density of 0.5 and 1.0 mmol m?2 S?1 and with three nutrient regimes until the third trifoliolate leaf of soybean or the sixth leaf of sugar beet had finished expanding. Net rates of CO2 exchange of the most recently expanded leaves were then measured at both 350 and 700 μl 1?1 CO2. Plants grown at the elevated CO2 concentration had net rates of leaf CO2 exchange which were reduced by 33% in sugar beet and 23% in soybean when measured at 350 μl 1?1 CO2 and when averaged over all treatments. Negative photosynthetic adjustment to elevated CO2 concentration was not greater at 20 than at 25°C, was not greater at a photon flux density of 1.0 than at 0.5 mmol m?2 S?1 and was not greater with limiting nutrients. Furthermore, in soybean, negative photosynthetic adjustment could be induced by a single night at elevated CO2 concentration, with net rates of CO2 exchange the next day equal to those of leaves of plants grown from seed at the elevated concentration of CO2. These patterns do not support either the feedback-inhibition or the nutrient-stress hypothesis of photosynthetic adjustment to elevated concentrations of CO2. 相似文献
40.
Protein synthesis in dormant embryos of sugar maple ( Acer saccharum ) was investigated in seeds stratified at 4°C or incubated at 15°C. Seeds stratified at 4°C germinated after 27 days; seeds incubated at 15°C failed to germinate. Stratification increased the embryo's capacity for protein synthesis by day 11 as measured by in vivo incorporation of [35 S]-methionine into purified protein. At 4°C protein synthesis in the embryonic axis rose in a linear fashion prior to germination, whereas in cotyledons it increased until day 20 and then declined. Analysis of radiolabelled proteins by two-dimensional gel electrophoresis revealed that the levels of specific proteins were altered by temperature, primarily in the cotyledons. Several proteins were expressed in the cotyledons at 15°C but were absent in unstratified embryos and in embryos stratified at 4°C. That is, the expression of these proteins was repressed during stratification and release from dormancy. Levels of other proteins in the cotyledons declined at 4°C during stratification. We suggest that one or more of these proteins may be associated with the inhibition of growth of the embryonic axis imposed by the cotyledons. 相似文献