全文获取类型
收费全文 | 10163篇 |
免费 | 669篇 |
国内免费 | 2篇 |
出版年
2023年 | 64篇 |
2022年 | 148篇 |
2021年 | 268篇 |
2020年 | 141篇 |
2019年 | 201篇 |
2018年 | 232篇 |
2017年 | 223篇 |
2016年 | 355篇 |
2015年 | 522篇 |
2014年 | 585篇 |
2013年 | 732篇 |
2012年 | 790篇 |
2011年 | 767篇 |
2010年 | 544篇 |
2009年 | 413篇 |
2008年 | 572篇 |
2007年 | 534篇 |
2006年 | 558篇 |
2005年 | 475篇 |
2004年 | 489篇 |
2003年 | 421篇 |
2002年 | 402篇 |
2001年 | 135篇 |
2000年 | 86篇 |
1999年 | 107篇 |
1998年 | 93篇 |
1997年 | 94篇 |
1996年 | 67篇 |
1995年 | 41篇 |
1994年 | 52篇 |
1993年 | 46篇 |
1992年 | 49篇 |
1991年 | 44篇 |
1990年 | 38篇 |
1989年 | 38篇 |
1988年 | 34篇 |
1987年 | 31篇 |
1986年 | 29篇 |
1985年 | 35篇 |
1984年 | 35篇 |
1983年 | 32篇 |
1982年 | 38篇 |
1981年 | 32篇 |
1980年 | 31篇 |
1979年 | 21篇 |
1977年 | 16篇 |
1976年 | 15篇 |
1975年 | 15篇 |
1974年 | 13篇 |
1973年 | 20篇 |
排序方式: 共有10000条查询结果,搜索用时 15 毫秒
691.
We develop a maximum penalized-likelihood (MPL) method to estimate the fitnesses of amino acids and the distribution of selection coefficients (S = 2Ns) in protein-coding genes from phylogenetic data. This improves on a previous maximum-likelihood method. Various penalty functions are used to penalize extreme estimates of the fitnesses, thus correcting overfitting by the previous method. Using a combination of computer simulation and real data analysis, we evaluate the effect of the various penalties on the estimation of the fitnesses and the distribution of S. We show the new method regularizes the estimates of the fitnesses for small, relatively uninformative data sets, but it can still recover the large proportion of deleterious mutations when present in simulated data. Computer simulations indicate that as the number of taxa in the phylogeny or the level of sequence divergence increases, the distribution of S can be more accurately estimated. Furthermore, the strength of the penalty can be varied to study how informative a particular data set is about the distribution of S. We analyze three protein-coding genes (the chloroplast rubisco protein, mammal mitochondrial proteins, and an influenza virus polymerase) and show the new method recovers a large proportion of deleterious mutations in these data, even under strong penalties, confirming the distribution of S is bimodal in these real data. We recommend the use of the new MPL approach for the estimation of the distribution of S in species phylogenies of protein-coding genes. 相似文献
692.
Alessandra V.R. da Silva Bibiana M. De Souza Marcia P. dos Santos Cabrera Nathalia B. Dias Paulo C. Gomes João Ruggiero Neto Rodrigo G. Stabeli Mario S. Palma 《生物化学与生物物理学报:生物膜》2014
Polycationic peptides may present their C-termini in either amidated or acidic form; however, the effects of these conformations on the mechanisms of interaction with the membranes in general were not properly investigated up to now. Protonectarina-MP mastoparan with an either amidated or acidic C-terminus was utilized to study their interactions with anionic and zwitterionic vesicles, using measurements of dye leakage and a combination of H/D exchange and mass spectrometry to monitor peptide–membrane interactions. Mast cell degranulation, hemolysis and antibiosis assays were also performed using these peptides, and the results were correlated with the structural properties of the peptides. The C-terminal amidation promotes the stabilization of the secondary structure of the peptide, with a relatively high content of helical conformations, permitting a deeper interaction with the phospholipid constituents of animal and bacterial cell membranes. The results suggested that at low concentrations Protonectarina-MP interacts with the membranes in a way that both terminal regions remain positioned outside the external surface of the membrane, while the α-carbon backbone becomes partially embedded in the membrane core and changing constantly the conformation, and causing membrane destabilization. The amidation of the C-terminal residue appears to be responsible for the stabilization of the peptide conformation in a secondary structure that is richer in α-helix content than its acidic congener. The helical, amphipathic conformation, in turn, allows a deeper peptide–membrane interaction, favoring both biological activities that depend on peptide structure recognition by the GPCRs (such as exocytosis) and those activities dependent on membrane perturbation (such as hemolysis and antibiosis). 相似文献
693.
Mario Cappiello Roberta MoschiniFrancesco Balestri Umberto MuraAntonella Del-Corso 《Biochemical and biophysical research communications》2014
The possible preferential action exerted by an inhibitor on the transformation of one of two agonist substrates catalyzed by the same enzyme has recently been reported in studies on aldose reductase inhibition. This event was defined as “intra-site differential inhibition” and the molecules able to exert this action as “differential inhibitors”. This work presents some basic kinetic models describing differential inhibition. Using a simple analytic approach, the results show that differential inhibition can occur through either competitive or mixed type inhibition in which the inhibitor prevalently targets the free enzyme. The results may help in selecting molecules whose differential inhibitory action could be advantageous in controlling the activity of enzymes acting on more than one substrate. 相似文献
694.
Per-Arne Svensson Björn Wahlstrand Maja Olsson Philippe Froguel Mario Falchi Richard N. Bergman Philip G. McTernan Thomas Hedner Lena M.S. Carlsson Peter Jacobson 《Biochemical and biophysical research communications》2014
Risk alleles within a gene desert at the 9p21 locus constitute the most prevalent genetic determinant of cardiovascular disease. Previous research has demonstrated that 9p21 risk variants influence gene expression in vascular tissues, yet the biological mechanisms by which this would mediate atherosclerosis merits further investigation. To investigate possible influences of this locus on other tissues, we explored expression patterns of 9p21-regulated genes in a panel of multiple human tissues and found that the tumor suppressor CDKN2B was highly expressed in subcutaneous adipose tissue (SAT). CDKN2B expression was regulated by obesity status, and this effect was stronger in carriers of 9p21 risk alleles. Covariation between expression of CDKN2B and genes implemented in adipogenesis was consistent with an inhibitory effect of CDKN2B on SAT proliferation. Moreover, studies of postprandial triacylglycerol clearance indicated that CDKN2B is involved in down-regulation of SAT fatty acid trafficking. CDKN2B expression in SAT correlated with indicators of ectopic fat accumulation, including markers of hepatic steatosis. Among genes regulated by 9p21 risk variants, CDKN2B appears to play a significant role in the regulation of SAT expandability, which is a strong determinant of lipotoxicity and therefore might contribute to the development of atherosclerosis. 相似文献
695.
Airoldi C Zona C Sironi E Colombo L Messa M Aurilia D Gregori M Masserini M Salmona M Nicotra F La Ferla B 《Journal of biotechnology》2010,156(4):317-324
Curcumin derivatives with high chemical stability, improved solubility and carrying a functionalized appendage for the linkage to other entities, have been synthesized in a straightforward manner. All compounds retained Curcumin ability to bind Aβ peptide oligomers without inducing their aggregation. Moreover all Curcumin derivatives were able to stain very efficiently Aβ deposits. 相似文献
696.
Fernanda A. H. Batista Leandro S. Goto Wanius Garcia Derminda I. de Moraes Mario de Oliveira Neto Igor Polikarpov Marcia R. Cominetti Heloísa S. Selistre-de-Araújo Leila M. Beltramini Ana Paula Ulian Araújo 《European biophysics journal : EBJ》2010,39(8):1193-1205
Lectins have been classified into a structurally diverse group of proteins that bind carbohydrates and glycoconjugates with
high specificity. They are extremely useful molecules in the characterization of saccharides, as drug delivery mediators,
and even as cellular surface makers. In this study, we present camptosemin, a new lectin from Camptosema ellipticum. It was characterized as an N-acetyl-d-galactosamine-binding homo-tetrameric lectin, with a molecular weight around 26 kDa/monomers. The monomers were stable over
a wide range of pH values and exhibited pH-dependent oligomerization. Camptosemin promoted adhesion of breast cancer cells
and hemagglutination, and both activities were inhibited by its binding of sugar. The stability and unfolding/folding behavior
of this lectin was characterized using fluorescence and far-UV circular dichroism spectroscopies. The results indicate that
chemical unfolding of camptosemin proceeds as a two-state monomer-tetramer process. In addition, small-angle X-ray scattering
shows that camptosemin behaves as a soluble and stable homo-tetramer molecule in solution. 相似文献
697.
A. Palamidessi I. Testa E. Frittoli S. Barozzi M. Garrè D. Mazza P. P. Di Fiore A. Diaspro G. Scita Mario Faretta 《European biophysics journal : EBJ》2010,39(6):947-957
The dissection of the molecular circuitries at the base of cell life and the identification of their abnormal transformation
during carcinogenesis rely on the characterization of biological phenotypes generated by targeted overexpression or deletion
of gene products through genetic manipulation. Fluorescence microscopy provides a wide variety of tools to monitor cell life
with minimal perturbations. The observation of living cells requires the selection of a correct balance between temporal,
spatial and “statistical” resolution according to the process to be analyzed. In the following paper ad hoc developed optical
tools for dynamical tracking from cellular to molecular resolution will be presented. Particular emphasis will be devoted
to discuss how to exploit light–matter interaction to selectively target specific molecular species, understanding the relationships
between their intracellular compartmentalization and function. 相似文献
698.
Robert F. Hennigan Lauren A. Foster Mary F. Chaiken Timmy Mani Michelle M. Gomes Andrew B. Herr Wallace Ip 《Molecular and cellular biology》2010,30(1):54-67
Neurofibromatosis type 2 is an inherited autosomal disorder caused by biallelic inactivation of the NF2 tumor suppressor gene. The NF2 gene encodes a 70-kDa protein, merlin, which is a member of the ezrin-radixin-moesin (ERM) family. ERM proteins are believed to be regulated by a transition between a closed conformation, formed by binding of their N-terminal FERM domain and C-terminal tail domain (CTD), and an open conformation, in which the two domains do not interact. Previous work suggests that the tumor suppressor function of merlin is similarly regulated and that only the closed form is active. Therefore, understanding the mechanisms that control its conformation is crucial. We have developed a series of probes that measures merlin conformation by fluorescence resonance energy transfer, both as purified protein and in live cells. Using these tools, we find that merlin exists predominately as a monomer in a stable, closed conformation that is mediated by the central α-helical domain. The contribution from the FERM-CTD interaction to the closed conformation appears to be less important. Upon phosphorylation or interaction with an effector protein, merlin undergoes a subtle conformational change, suggesting a novel mechanism that modulates the interaction between the FERM domain and the CTD.Neurofibromatosis type 2 is an inherited autosomal disorder that is characterized by bilateral schwannomas of the eighth cranial nerve. The tumor suppressor gene responsible for this disorder, NF2, was cloned in 1993 (45). Biallelic inactivation of the NF2 gene is also seen in spontaneous schwannoma, meningioma, and malignant mesothelioma (22). In mouse models, deletion of the Nf2 gene is embryonic lethal, indicating an essential role for NF2 in development (24). Heterozygous mice develop a variety of aggressive metastatic tumors that have lost the wild-type allele (23). Targeted deletion of the Nf2 gene in Schwann cells leads to schwannoma formation (7). In vitro, Nf2-null cells grow to significantly higher densities (31), suggesting that contact inhibition of growth is impaired in these cells and that mediation of growth arrest at high cell density may be the basis for the tumor suppressor function of the NF2 gene. In normal fibroblasts, merlin is inactive as a growth suppressor in subconfluent cells, becoming activated as they approach confluence, thereby effecting contact inhibition of growth (26).The NF2 gene encodes a 70-kDa protein called merlin (for moesin, ezrin, radixin-like protein), which shares significant homology with members of the ezrin-radixin-moesin (ERM) branch of the Band 4.1 superfamily (45). The domain structure of merlin, also shared with other ERM proteins, consists of an N-terminal FERM domain, followed by a central α-helical region (CH) and a C-terminal tail domain (CTD). The merlin FERM domain has relatively high sequence similarity with other ERM family members, a 60 to 70% identity over the first 300 amino acids. The CH domain and the CTD show much lower identity (28 to 36%); however, the α-helical character of the CH domain is preserved, as is the heptad repeat pattern typical of α-helices that form coiled coils (46).The critical point of regulation of all the ERM proteins is a high-affinity intramolecular interaction between the C-terminal domain and the FERM domain (4) (Fig. (Fig.1).1). The FERM domain folds into a three-lobed cloverleaf structure that acts as a multifaceted docking site for protein binding partners (16, 39). The CTD, consisting of four major and two minor helices and a beta sheet, binds to the FERM domain by extending across the face of the F2 and F3 lobes (32). This intramolecular head-to-tail binding results in a “closed” conformation, with the C-terminal domain covering much of the surface of the FERM domain (32, 44). For ezrin, radixin and moesin, the CTD functions as a mask, blocking access of effector molecules, such as the cell surface receptors CD44 and ICAM2 and adaptor molecules, like EBP50/NHERF, to sites on the surface of the FERM domain (11, 25, 44). The interaction between the CTD and FERM domain is regulated by phosphatidyl inositol-(4,5)-bisphosphate (PIP2) binding to the FERM domain and by phosphorylation of a critical residue in the CTD (3, 6, 10, 49). This residue, threonine 567 in ezrin, is conserved throughout the ERM family (21). Phosphorylation introduces a negative charge and a bulky side group that effectively reduces the affinity of the interaction, releasing the CTD from the FERM domain and causing a transition to an open conformation. Low-angle rotary shadowing electron microscopy (13) and biochemical studies (12) of purified radixin suggest that in the open conformation it is an extended filamentous structure with globular N and C termini that is greater than 240 Å in length. Signal transduction systems, such as the epidermal growth factor and Rho A pathways, induce phosphorylation of ERM proteins at the conserved C-terminal threonine via a number of kinases, including Rho kinase and protein kinase Cα (21, 28). Thus, conformational regulation of ERM proteins can be a point of integration of ERM activity with signal transduction pathways. The overall concept of ERM regulation, then, is centered upon a transition between an inactive, closed conformation that is mediated by the FERM-CTD interaction and an active, open conformation that is regulated by phosphorylation. In these two states, ERM proteins likely interact with different sets of binding partners, resulting in distinct functional outcomes.Open in a separate windowFIG. 1.ERM tertiary structure as represented by the crystal structure of full-length Sf-moesin (20), but with the merlin amino acid sequence substituted for Sf-moesin. Approximate boundary amino acid residues for all domains appear at the top of the figure. Each domain is assigned a different color. The ERM structure consists of an N-terminal FERM domain folded into three lobes, F1, F2, and F3. This is followed by a central α-helical domain containing three subhelices (αA, αB, and αC) and a CTD with four short helices. An ERM protein is thought to have an open conformation, an extended structure with the FERM domain and the CTD separated by the α-helical domain, that is more than 240 Å long. In the closed conformation, the α-helical domain bends at the αA-αB junction and again at the αB-αC junction, causing the CTD to be positioned over F2 and F3 of the FERM domain. More than half of the surface of the FERM domain is masked by interaction with the CTD, αA, and parts of αB and αC.Like the classical ERMs, merlin is also thought to be regulated by changes in conformation. The FERM domain and the CTD of merlin interact with each other, albeit at a lower level of affinity than the ezrin FERM domain and the CTD (29). There are important differences, however, between merlin and the other ERM proteins. First, phosphorylation of the conserved C-tail threonine T576 has not been reported to occur in mammalian merlin, and nonphosphorylatable and phosphomimetic substitutions at this site have no effect on merlin activity (42). Instead, merlin is phosphorylated at serine 518 in the CTD, a target of the p21-activated kinase PAK and protein kinase A (1, 18, 47). The growth-suppressive function of merlin is activated by dephosphorylation of S518 by the phosphatase PP1δ in a density-dependent manner (14). Second, it was reported in a study using FERM domain and CTD truncates of merlin that only cotransfection of both the N-and C-terminal halves resulted in growth suppression (38). Together, these observations suggested a model of inactive, phosphorylated merlin in an open conformation that, upon cell-to-cell contact, is dephosphorylated and transitions to a closed, growth suppressive conformation.The existing model for conformational regulation described above is inferred from indirect data and assays that generally measure the interaction of isolated FERM and CTD truncates rather than full-length molecules (9, 29, 38). It has been impossible to test directly because tools have not been available to specifically assay for either the open or the closed form of merlin. Therefore, we have developed a series of probes that measures merlin conformation by fluorescence resonance energy transfer (FRET), both as purified protein and in live cells. Using these tools, we show that merlin exists predominately as a monomer in a stable, largely closed conformation. Additionally, we find that the closed conformation is largely mediated by the central α-helical domain; the contribution of the FERM-CTD interaction appears to be less significant than previously thought. Finally, we find that phosphorylation and protein interaction cause unexpectedly small changes in merlin conformation. We propose a new and more refined model for merlin regulation, in which merlin function is regulated by specific but subtle conformational changes that modulate the interaction between the FERM domain and the CTD. 相似文献
699.
Mario Melletti M. M. Delgado Vincenzo Penteriani Marzia Mirabile Luigi Boitani 《Journal of Ethology》2010,28(3):421-428
Many animals aggregate into organized temporary or stable groups under the influence of biotic and abiotic factors, and some
studies have shown the influence of habitat features on animal aggregation. This study, conducted from 2002 to 2004 in the
Dzanga-Ndoki National Park, Central African Republic, studied a herd of forest buffaloes (Syncerus caffer nanus) to determine whether spatial aggregation patterns varied by season and habitat. Our results show that both habitat structure
and season influenced spatial aggregation patterns. In particular, in open habitats such as clearings, the group covered a
larger area when resting and was more rounded in shape compared to group properties noted in forest during the wet season.
Moreover, forest buffaloes had a more aggregated spatial distribution when resting in clearings than when in the forest, and
individual positions within the herd in the clearing habitat varied with age and sex. In the clearings, the adult male (n = 24) was generally, on most occasions, located in the centre of the herd (n = 20), and he was observed at the border only four times. In contrast, females (n = 80) occupied intermediate (n = 57), peripheral (n = 14) and central positions (n = 9) within the group. Juveniles (n = 77) also occurred in intermediate (n = 64) and peripheral positions (n = 13). Based on these results, we concluded that habitat characteristics and social behaviour can have relevant effects on
the spatial distribution of animals within a group. 相似文献
700.
Fabricio Fernandes Fontana Celso Tadeu Barbosa dos Santos Flavia Maria Esteves Ademir Rocha Geisa Ferreira Fernandes Cristiane Candida do Amaral Marcos Abel Domingues Zoilo Pires De Camargo Mario León Silva-Vergara 《Mycopathologia》2010,169(3):159-165
There is some evidence that dogs can be naturally infected by Paracoccidioides brasiliensis in endemic areas of paracoccidioidomycosis. In order to evaluate canine infection with this fungus, a survey with 149 urban
and 126 rural dogs was carried out using ELISA and intradermal tests with the gp43 antigen of P. brasiliensis in Uberaba, Minas Gerais state of Brazil. Forty-one out of 149 urban dogs were euthanatized and had their lungs, liver and
spleen removed. One slice from each viscera was processed for histopathological examination and the remaining was homogenized and then cultivated on mycobiotic agar
at room temperature and Fava-Netto medium at 35°C and observed for 12 weeks. Of urban dogs, 75 (50.3%) were small adult females,
56 (36%) were strays, while 93 (64%) had been donated to the municipal zoonosis control center. Nine (6.2%) had a positive
intradermal test without statistical differences regarding gender, race, nutritional status or origin. No colonies with microscopic
or morphology appearances resembling P. brasiliensis were isolated, nor granulomatous process or fungal structures were observed from histopathological examination. Eighty (53.6%)
of the urban dogs presented seroreactivity, without statistical differences regarding gender, race, nutritional state, origin,
or positive intradermal test. Of 126 rural dogs, 102 (80.5%) presented antibodies against gp43 antigen, and this was statistically
significant in relation to the reactivity detected in urban dogs (P = 0.0001). Thus, dogs are commonly infected with P. brasiliensis, but they probably present natural resistance to develop paracoccidioidomycosis. 相似文献