全文获取类型
收费全文 | 651篇 |
免费 | 86篇 |
国内免费 | 4篇 |
出版年
2021年 | 8篇 |
2020年 | 6篇 |
2019年 | 5篇 |
2018年 | 8篇 |
2017年 | 5篇 |
2016年 | 12篇 |
2015年 | 18篇 |
2014年 | 20篇 |
2013年 | 31篇 |
2012年 | 28篇 |
2011年 | 37篇 |
2010年 | 25篇 |
2009年 | 21篇 |
2008年 | 19篇 |
2007年 | 34篇 |
2006年 | 19篇 |
2005年 | 30篇 |
2004年 | 20篇 |
2003年 | 24篇 |
2002年 | 20篇 |
2001年 | 17篇 |
2000年 | 16篇 |
1999年 | 16篇 |
1998年 | 16篇 |
1997年 | 7篇 |
1995年 | 7篇 |
1994年 | 8篇 |
1993年 | 16篇 |
1992年 | 28篇 |
1991年 | 21篇 |
1990年 | 10篇 |
1989年 | 9篇 |
1988年 | 14篇 |
1987年 | 11篇 |
1986年 | 12篇 |
1985年 | 9篇 |
1984年 | 18篇 |
1982年 | 5篇 |
1980年 | 10篇 |
1979年 | 10篇 |
1978年 | 9篇 |
1976年 | 6篇 |
1975年 | 5篇 |
1972年 | 11篇 |
1971年 | 7篇 |
1970年 | 3篇 |
1969年 | 3篇 |
1968年 | 4篇 |
1967年 | 4篇 |
1961年 | 4篇 |
排序方式: 共有741条查询结果,搜索用时 15 毫秒
61.
Long-chain hydroxy acid oxydase (HAO) is a member of a flavoenzyme family with significant amino acid sequence similarity and strongly conserved three-dimensional structure; in particular, active-site amino acids involved in catalysis are invariant, with one exception, and numerous enzymatic studies suggest an identical chemical mechanism involving an intermediate carbanion for all family members. Known physiological substrates are a variety of L-2-hydroxy acids. Peroxisomal HAO differs from the other family members in that its actual physiological substrate is not known; it was first described as an L-amino acid oxidase, and recently was identified as an enzyme that converts creatol (hydroxycreatinine) to methylguanidine (a metabolite involved in a variety of uremic syndromes). Creatol (2-amino-5-hydroxy-1-methyl-4(5H)imidazolone) is not a 2-hydroxy acid. We show in this work that 2-hydroxyphenyl acetohydroxamate (HYPAH, the hydroxamate of mandelic acid), a compound that bears similarity both to mandelate (one of the best substrates known) and to creatol, is turned over by HAO, but between 10- and 100-fold less efficiently than mandelate itself. The compound also binds to the active site of homologous flavocytochrome b(2) (L-lactate dehydrogenase). Comparative pH-rate studies for mandelate and its hydroxamate suggest that HYPAH may bind in its ionized form. Both pH-rate profiles are bell-shaped curves, as are those determined for two other family members, flavocytochrome b(2) and mandelate dehydrogenase; while the group with an acid pK(a) between 5 and 6 is most likely the active-site histidine (the residue which abstracts the substrate C2 proton), the identity of the basic group is less clear. It has been proposed to be one of the active site arginines (Lehoux, I., and Mitra, B. (1999) Biochemistry38, 5836-5848); we suggest as an alternative that it could be the lysine residue that interacts with the flavin N1 and O2 positions and stabilizes the negative charge of reduced flavin. In addition to these studies, we have found that HAO is competitively inhibited by benzohydroxamate, which is one atom shorter than HYPAH; its affinity is nearly 100-fold lower than that of the substrate, in contrast to the strong inhibition it exerts on mandelate racemase (Maurice, St. M., and Bearne, S. L. (2000) Biochemistry39, 13324-13335). In the latter case, the 100-fold higher affinity compared to mandelate was proposed to arise from the fact that the hydroxamate can mimic the enolic intermediate which lies on the reaction pathway after C2 proton abstraction. Thus our results do not support the existence of a similar enolic intermediate for HAO (and probably its homologues), although they do not disprove it. 相似文献
62.
Rodríguez-Pachón JM Martín H North G Rotger R Nombela C Molina M 《The Journal of biological chemistry》2002,277(30):27094-27102
Modulation of host cellular GTPases through the injection of the effector proteins SopE2 and SptP is essential for Salmonella typhimurium to enter into non-phagocytic cells. Here we show that expression of the guanine nucleotide exchange factor for Cdc42 SopE2 in Saccharomyces cerevisiae leads to the activation of Fus3 and Kss1 MAPKs, which operate in the mating and filamentation pathways, causing filamentous growth in haploid yeast cells. Furthermore, it promotes the activation of the cell integrity MAPK Slt2. Cdc42 activation by removal of its putative intrinsic GTPase-activating proteins (GAPs), Rga1, Rga2, and Bem3, also results in the phosphorylation of Kss1, Fus3, and Slt2 MAPKs. These data support the role of these GAP proteins as negative regulators of Cdc42, confirm the modulating effect of this GTPase on the filamentation and mating pathways and point to a novel connection between Cdc42 and the cell integrity pathway. Cdc42-induced activation of Slt2 occurs in a mating and filamentation pathway-dependent manner, but it does not require the function of Rho1, which is the GTPase that operates in the cell integrity pathway. Moreover, we report that Salmonella SptP can act as a GAP for Cdc42 in S. cerevisiae, down-regulating MAPK-mediated signaling. Thus, yeast provides a useful system to study the interaction of bacterial pathogenic proteins with eukaryotic signaling pathways. Furthermore, these proteins can be used as a tool to gain insight into the mechanisms that regulate MAPK-mediated signaling in eukaryotes. 相似文献
63.
The initial rate of Fe(2+) movement across the inner envelope membrane of pea (Pisum sativum) chloroplasts was directly measured by stopped-flow spectrofluorometry using membrane vesicles loaded with the Fe(2+)-sensitive fluorophore, Phen Green SK. The rate of Fe(2+) transport was rapid, coming to equilibrium within 3s. The maximal rate and concentration dependence of Fe(2+) transport in predominantly right-side-out vesicles were nearly equivalent to those measured in largely inside-out vesicles. Fe(2+) transport was stimulated by an inwardly directed electrochemical proton gradient across right-side-out vesicles, an effect that was diminished by the addition of valinomycin in the presence of K(+). Fe(2+) transport was inhibited by Zn(2+), in a competitive manner, as well as by Cu(2+) and Mn(2+). These results indicate that inward-directed Fe(2+) transport across the chloroplast inner envelope occurs by a potential-stimulated uniport mechanism. 相似文献
64.
Use of Leu3a/3b for the accurate determination of CD4 subsets for measurement of intracellular cytokines 总被引:2,自引:0,他引:2
BACKGROUND: Identification of human T-helper cell subsets is possible by measurement of intracellular cytokines after coincubation of lymphocytes with phorbol myristate acetate (PMA), calcium ionophore, and brefeldin A for up to 20 h. However, exposure to PMA leads to internalization of membrane CD4 and to loss of resolution of the CD4+ cells. Detection of CD3+CD8- cells or preselection of CD4+ cells prior to stimulation is more cumbersome than direct measurement of CD4+ cells. We report the use of the Leu3a/Leu3b multiclone for the accurate determination of CD4 cells after PMA stimulation. METHODS: Peripheral blood lymphocytes were isolated from healthy normal donors and the proportion of CD3+ / CD4+ T cells was determined by flow cytometry before and after incubation with PMA, calcium ionophore, and brefeldin A for 20 h using a variety of anti-CD4 monoclonal antibodies. RESULTS: The Leu3a/3b multiclone reagent was the only anti-CD4 monoclonal antibody capable of resolving more than 98% of the initial CD4+ events after incubation with PMA. CONCLUSIONS: The higher signal-to-noise ratio associated with Leu3a3b reagent, compared with other CD4-specific antibodies available, allows the direct and accurate identification of the CD4 subset even after PMA treatment of cells. 相似文献
65.
Solution structure of the sixth LDL-A module of the LDL receptor 总被引:5,自引:0,他引:5
The low-density lipoprotein receptor (LDLR) is the primary mechanism for uptake of plasma cholesterol into cells and serves as a prototype for an entire class of cell surface receptors. The amino-terminal domain of the receptor consists of seven LDL-A modules; the third through the seventh modules all contribute to the binding of low-density lipoproteins (LDLs). Here, we present the NMR solution structure of the sixth LDL-A module (LR6) from the ligand binding domain of the LDLR. This module, which has little recognizable secondary structure, retains the essential structural features observed in the crystal structure of LDL-A module five (LR5) of the LDLR. Three disulfide bonds, a pair of buried residues forming a hydrophobic "mini-core", and a calcium-binding site that serves to organize the C-terminal lobe of the module all occupy positions in LR6 similar to those observed in LR5. The striking presence of a conserved patch of negative surface electrostatic potential among LDL-A modules of known structure suggests that ligand recognition by these repeats is likely to be mediated in part by electrostatic complementarity of receptor and ligand. Two variants of LR6, identified originally as familial hypercholesterolemia (FH) mutations, have been investigated for their ability to form native disulfide bonds under conditions that permit disulfide exchange. The first, E219K, lies near the amino-terminal end of LR6, whereas the second, D245E, alters one of the aspartate side chains that directly coordinate the bound calcium ion. After equilibration at physiologic calcium concentrations, neither E219K nor D245E folds to a unique disulfide isomer, indicating that FH mutations both within and distant from the calcium-binding site give rise to protein-folding defects. 相似文献
66.
Thilo Sascha Lange Michael Ezrokhi Anton V. Borovjagin Rafael Rivera-Len Melanie T. North Susan A. Gerbi 《Molecular biology of the cell》1998,9(10):2973-2985
The Nucleolar Localization Elements (NoLEs) of Xenopus laevis U3 small nucleolar RNA (snoRNA) have been defined. Fluorescein-labeled wild-type U3 snoRNA injected into Xenopus oocyte nuclei localized specifically to nucleoli as shown by fluorescence microscopy. Injection of mutated U3 snoRNA revealed that the 5′ region containing Boxes A and A′, known to be important for rRNA processing, is not essential for nucleolar localization. Nucleolar localization of U3 snoRNA was independent of the presence and nature of the 5′ cap and the terminal stem. In contrast, Boxes C and D, common to the Box C/D snoRNA family, are critical elements for U3 localization. Mutation of the hinge region, Box B, or Box C′ led to reduced U3 nucleolar localization. Results of competition experiments suggested that Boxes C and D act in a cooperative manner. It is proposed that Box B facilitates U3 snoRNA nucleolar localization by the primary NoLEs (Boxes C and D), with the hinge region of U3 subsequently base pairing to the external transcribed spacer of pre-rRNA, thus positioning U3 snoRNA for its roles in rRNA processing. 相似文献
67.
68.
Iwan G. Jones Michael North 《International journal of peptide research and therapeutics》1998,5(2-3):171-173
Summary The desymmetrisation ofendo-norborn-5-ene-2,3-dicarboxylic anhydride by proline esters has been used to prepare conformationally constrained pseudo-peptides
with two peptide chains parallel to one another. A Curtius rearrangement on the desymmetrication adduct produced the corresponding
isocyanate which was used to prepare both a peptide incorporating anendo-2-amino-3-carboxy-norborn-5-ene unit, and a pseudo-peptide with two peptide chains parallel to one another but offset by
the presence of a urea unit. The conformational analysis of the resulting peptides was carried out, and the norbornene unit
was found to induce the formation of β-turns and parallel β-sheets. 相似文献
69.
70.
Caryn S. Ross-Innes Irene Debiram-Beecham Maria O'Donovan Elaine Walker Sibu Varghese Pierre Lao-Sirieix Laurence Lovat Michael Griffin Krish Ragunath Rehan Haidry Sarmed S. Sami Philip Kaye Marco Novelli Babett Disep Richard Ostler Benoit Aigret Bernard V. North Pradeep Bhandari Adam Haycock Danielle Morris Stephen Attwood Anjan Dhar Colin Rees Matthew D. D. Rutter Peter D. Sasieni Rebecca C. Fitzgerald 《PLoS medicine》2015,12(1)
BackgroundBarrett''s esophagus (BE) is a commonly undiagnosed condition that predisposes to esophageal adenocarcinoma. Routine endoscopic screening for BE is not recommended because of the burden this would impose on the health care system. The objective of this study was to determine whether a novel approach using a minimally invasive cell sampling device, the Cytosponge, coupled with immunohistochemical staining for the biomarker Trefoil Factor 3 (TFF3), could be used to identify patients who warrant endoscopy to diagnose BE.ConclusionsThe Cytosponge-TFF3 test is safe and acceptable, and has accuracy comparable to other screening tests. This test may be a simple and inexpensive approach to identify patients with reflux symptoms who warrant endoscopy to diagnose BE. 相似文献