全文获取类型
收费全文 | 6991篇 |
免费 | 579篇 |
国内免费 | 523篇 |
出版年
2023年 | 95篇 |
2022年 | 94篇 |
2021年 | 135篇 |
2020年 | 166篇 |
2019年 | 238篇 |
2018年 | 248篇 |
2017年 | 203篇 |
2016年 | 193篇 |
2015年 | 243篇 |
2014年 | 346篇 |
2013年 | 498篇 |
2012年 | 245篇 |
2011年 | 324篇 |
2010年 | 257篇 |
2009年 | 314篇 |
2008年 | 296篇 |
2007年 | 298篇 |
2006年 | 284篇 |
2005年 | 273篇 |
2004年 | 234篇 |
2003年 | 200篇 |
2002年 | 187篇 |
2001年 | 175篇 |
2000年 | 179篇 |
1999年 | 164篇 |
1998年 | 164篇 |
1997年 | 141篇 |
1996年 | 128篇 |
1995年 | 131篇 |
1994年 | 122篇 |
1993年 | 121篇 |
1992年 | 110篇 |
1991年 | 104篇 |
1990年 | 107篇 |
1989年 | 84篇 |
1988年 | 99篇 |
1987年 | 86篇 |
1986年 | 65篇 |
1985年 | 101篇 |
1984年 | 109篇 |
1983年 | 76篇 |
1982年 | 76篇 |
1981年 | 68篇 |
1980年 | 57篇 |
1979年 | 39篇 |
1978年 | 39篇 |
1977年 | 39篇 |
1975年 | 28篇 |
1974年 | 28篇 |
1973年 | 30篇 |
排序方式: 共有8093条查询结果,搜索用时 33 毫秒
891.
Maria Sentandreu Pablo Leivar Guiomar Martín Elena Monte 《Plant signaling & behavior》2012,7(4):510-513
Plants need to accurately adjust their development after germination in the underground darkness to ensure survival of the seedling, both in the dark and in the light upon reaching the soil surface. Recent studies have established that the photoreceptors phytochromes and the bHLH phytochrome interacting factors PIFs regulate seedling development to adjust it to the prevailing light environment during post-germinative growth. However, complete understanding of the downstream regulatory network implementing these developmental responses is still lacking. In a recent work, published in The Plant Cell, we report a subset of PIF3-regulated genes in dark-grown seedlings that we have named MIDAs (MISREGULATED IN DARK). Analysis of their functional relevance using mutants showed that four of them present phenotypic alterations in the dark, and that each affected a particular facet of seedling development, suggesting organ-specific branching in the signal that PIF3 relays downstream. Furthermore, our results also showed an altered response to light in seedlings with an impaired PIF3/MIDA regulatory network, indicating that these factors might also be essential to initiate and optimize the developmental adjustment of the seedling to the light environment. 相似文献
892.
禽流感病毒H5N1 NS1蛋白是一种非结构蛋白,在病毒感染过程中发挥着重要的作用.构建基因截短的重组蛋白,可为进一步研究NS1不同结构域与宿主蛋白间的相互作用奠定基础.在成功克隆禽流感病毒H5N1全长NS1基因并测序的基础上,将部分截短基因序列克隆到表达栽体pET28a(+)上,构建基因截短的重组表达质粒pET28a-NS1-RBD和pET28a-NS1-ED,转化大肠埃希菌BL21(DE3),阳性重组质粒经IPTG诱导表达后进行SDS-PAGE检测,获得预期蛋白的表达,然后利用Ni-NTA树脂蛋白纯化系统对重组蛋白进行纯化,并通过Western Blotting进一步确认NS1及截短体蛋白的表达.结果表明,实验成功构建禽流感病毒H5N1亚型的NS1蛋白截短体,并在大肠埃希菌中高效表达,这为进一步研究NS1蛋白不同结构域与宿主蛋白的相互作用提供了实验材料,为深入研究NS1蛋白的生物学功能奠定了坚实基础. 相似文献
893.
894.
895.
Govada L Carpenter L da Fonseca PC Helliwell JR Rizkallah P Flashman E Chayen NE Redwood C Squire JM 《Journal of molecular biology》2008,378(2):387-397
C-protein is a major component of skeletal and cardiac muscle thick filaments. Mutations in the gene encoding cardiac C-protein [cardiac myosin binding protein-C (cMyBP-C)] are one of the principal causes of hypertrophic cardiomyopathy. cMyBP-C is a string of globular domains including eight immunoglobulin-like and three fibronectin-like domains termed C0-C10. It binds to myosin and titin, and probably to actin, and may have both a structural and a regulatory role in muscle function. To help to understand the pathology of the known mutations, we have solved the structure of the immunoglobulin-like C1 domain of MyBP-C by X-ray crystallography to a resolution of 1.55 Å. Mutations associated with hypertrophic cardiomyopathy are clustered at one end towards the C-terminus, close to the important C1C2 linker, where they alter the structural integrity of this region and its interactions. 相似文献
896.
Xu Y Nakajima Y Ito K Zheng H Oyama H Heiser U Hoffmann T Gärtner UT Demuth HU Yoshimoto T 《Journal of molecular biology》2008,375(3):708-719
A new inhibitor, H-Ala-Ile-pyrrolidin-2-yl boronic acid, was developed as an inhibitor against prolyl tripeptidyl aminopeptidase with a Ki value of 88.1 nM. The structure of the prolyl tripeptidyl aminopeptidase complexed with the inhibitor (enzyme-inhibitor complex) was determined at 2.2 Å resolution. The inhibitor was bound to the active site through a covalent bond between Ser603 and the boron atom of the inhibitor. This structure should closely mimic the structure of the reaction intermediate between the enzyme and substrate. We previously proposed that two glutamate residues, Glu205 and Glu636, are involved in the recognition of substrates. In order to clarify the function of these glutamate residues in substrate recognition, three mutant enzymes, E205A, E205Q, and E636A were generated by site-directed mutagenesis. The E205A mutant was expressed as an inclusion body. The E205Q mutant was expressed in soluble form, but no activity was detected. Here, the structures of the E636A mutant and its complex with the inhibitor were determined. The inhibitor was located at almost the same position as in the wild-type enzyme-inhibitor complex. The amino group of the inhibitor interacted with Glu205 and the main-chain carbonyl group of Gln203. In addition, a water molecule in the place of Glu636 of the wild-type enzyme interacted with the amino group of the inhibitor. This water molecule was located near the position of Glu636 in the wild-type and formed a hydrogen bond with Gln203. The kcat/KM values of the E636A mutant toward the two substrates used were smaller than those of the wild-type by two orders of magnitude. The Ki value of our inhibitor for the E636A mutant was 48.8 μM, which was 554-fold higher than that against the wild-type enzyme. Consequently, it was concluded that Glu205 and Glu636 are significant residues for the N-terminal recognition of a substrate. 相似文献
897.
Prevention of abnormal misfolding and aggregation of α synuclein (syn) protein in vulnerable neurons should be viable therapeutic strategies for reducing pathogenesis in Parkinson's disease. The nonamyloid component (NAC) region of α-syn shows strong tendencies to form β-sheet structures, and deletion of this region has been shown to reduce aggregation and toxicity in vitro and in vivo. The binding of a molecular species to this region may mimic the effects of such deletions. Single-chain variable fragment (scFv) antibodies retain the binding specificity of antibodies and, when genetically manipulated to create high-diversity libraries, allow in vitro selection against peptides. Accordingly, we used a yeast surface display library of an entire naïve repertoire of human scFv antibodies to select for binding to a NAC peptide. Candidate scFv antibodies (after transfer to mammalian expression vectors) were screened for viability in a neuronal cell line by transient cotransfection with A53T mutant α-syn. This provided a ranking of the protective efficacies of the initial panel of intracellular antibodies (intrabodies). High steady-state expression levels and apparent conformational epitope binding appeared more important than in vitro affinity in these assays. None of the scFv antibodies selected matched the sequences of previously reported anti-α-syn scFv antibodies. A stable cell line expressing the most effective intrabody, NAC32, showed highly significant reductions in abnormal aggregation in two separate models. Recently, intrabodies have shown promising antiaggregation and neuroprotective effects against misfolded mutant huntingtin protein. The NAC32 study extends such work significantly by utilizing information about the pathogenic capacity of a specific α-syn region to offer a new generation of in vitro-derived antibody fragments, both for further engineering as direct therapeutics and as a tool for rational drug design for Parkinson's disease. 相似文献
898.
Beta-lactoglobulin assembles into amyloid through sequential aggregated intermediates 总被引:1,自引:0,他引:1
We have investigated the aggregation and amyloid fibril formation of bovine β-lactoglobulin variant A, with a focus on the early stages of aggregation. We used noncovalent labeling with thioflavin T and 1-anilino-8-naphthalenesulfonate to follow the conformational changes occurring in β-lactoglobulin during aggregation using time resolved luminescence. 1-Anilino-8-naphthalenesulfonate monitored the involvement of the hydrophobic core/calyx of β-lactoglobulin in the aggregation process. Thioflavin T luminescence monitored the formation of amyloid. The luminescence lifetime distributions of both probes showed changes that could be attributed to conformational changes occurring during and following aggregation. To correlate the luminescence measurements with the degree of aggregation and the morphology of the aggregates, we also measured dynamic light scattering and atomic force microscopy images. We evaluated the relative stability of the intermediates with an assay that is sensitive to aggregation reversibility. Our results suggest that initial aggregation during the first 5 days occurred with partial disruption of the characteristic calyx in β-lactoglobulin. As the globular aggregates grew from days 5 to 16, the calyx was completely disrupted and the globular aggregates became more stable. After this second phase of aggregation, conversion into a fibrillar form occurred, marking the growth phase, and still more changes in the luminescence signals were observed. Based on these observations, we propose a three-step process by which monomer is converted first into weakly associated aggregates, which rearrange into stable aggregates, which eventually convert into protofibrils that elongate in the growth phase. 相似文献
899.
Cornilescu G Ulijasz AT Cornilescu CC Markley JL Vierstra RD 《Journal of molecular biology》2008,383(2):403-413
The unique photochromic absorption behavior of phytochromes (Phys) depends on numerous reversible interactions between the bilin chromophore and the associated polypeptide. To help define these dynamic interactions, we determined by NMR spectroscopy the first solution structure of the chromophore-binding cGMP phosphodiesterase/adenylcyclase/FhlA (GAF) domain from a cyanobacterial Phy assembled with phycocyanobilin (PCB). The three-dimensional NMR structure of Synechococcus OS-B′ cyanobacterial Phy 1 in the red-light-absorbing state of Phy (Pr) revealed that PCB is bound to Cys138 of the GAF domain via the A-ring ethylidene side chain and is buried within the GAF domain in a ZZZsyn,syn,anti configuration. The D ring of the chromophore sits within a hydrophobic pocket and is tilted by approximately 80° relative to the B/C rings by contacts with Lys52 and His169. The solution structure revealed remarkable flexibility for PCB and several adjacent amino acids, indicating that the Pr chromophore has more freedom in the binding pocket than anticipated. The propionic acid side chains of rings B and C and Arg101 and Arg133 nearby are especially mobile and can assume several distinct and energetically favorable conformations. Mutagenic studies on these arginines, which are conserved within the Phy superfamily, revealed that they have opposing roles, with Arg101 and Arg133 helping stabilize and destabilize the far-red-light-absorbing state of Phy (Pfr), respectively. Given the fact that the Synechococcus OS-B′ GAF domain can, by itself, complete the Pr → Pfr photocycle, it should now be possible to determine the solution structure of the Pfr chromophore and surrounding pocket using this Pr structure as a framework. 相似文献
900.
Weiergräber OH Stangler T Thielmann Y Mohrlüder J Wiesehan K Willbold D 《Journal of molecular biology》2008,381(5):1320-1331
The γ-aminobutyric acid type A (GABAA) receptor-associated protein is a versatile adaptor protein playing an important role in intracellular vesicle trafficking, particularly in neuronal cells. We present the X-ray structure of the soluble form of human GABAA receptor-associated protein complexed with a high-affinity synthetic peptide at 1.3 Å resolution. The data shed light on the probable binding modes of key interaction partners, including the GABAA receptor and the cysteine protease Atg4. The resulting models provide a structural background for further investigation of the unique biological properties of this protein. 相似文献