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991.
Andrea F. Moon Geoffrey A. Mueller Xuejun Zhong Lars C. Pedersen 《Protein science : a publication of the Protein Society》2010,19(5):901-913
Protein crystallographers are often confronted with recalcitrant proteins not readily crystallizable, or which crystallize in problematic forms. A variety of techniques have been used to surmount such obstacles: crystallization using carrier proteins or antibody complexes, chemical modification, surface entropy reduction, proteolytic digestion, and additive screening. Here we present a synergistic approach for successful crystallization of proteins that do not form diffraction quality crystals using conventional methods. This approach combines favorable aspects of carrier‐driven crystallization with surface entropy reduction. We have generated a series of maltose binding protein (MBP) fusion constructs containing different surface mutations designed to reduce surface entropy and encourage crystal lattice formation. The MBP advantageously increases protein expression and solubility, and provides a streamlined purification protocol. Using this technique, we have successfully solved the structures of three unrelated proteins that were previously unattainable. This crystallization technique represents a valuable rescue strategy for protein structure solution when conventional methods fail. 相似文献
992.
Michael S. Malamas Keith Barnes Yu Hui Matthew Johnson Frank Lovering Jeff Condon William Fobare William Solvibile Jim Turner Yun Hu Eric S. Manas Kristi Fan Andrea Olland Rajiv Chopra Jonathan Bard Menelas N. Pangalos Peter Reinhart Albert J. Robichaud 《Bioorganic & medicinal chemistry letters》2010,20(7):2068-2073
The proteolytic enzyme β-secretase (BACE1) plays a central role in the synthesis of the pathogenic β-amyloid in Alzheimer’s disease. Recently, we reported small molecule acylguanidines as potent BACE1 inhibitors. However, many of these acylguanidines have a high polar surface area (e.g. as measured by the topological polar surface area or TPSA), which is unfavorable for crossing the blood–brain barrier. Herein, we describe the identification of the 2-aminopyridine moiety as a bioisosteric replacement of the acylguanidine moiety, which resulted in inhibitors with lower TPSA values and superior brain penetration. X-ray crystallographic studies indicated that the 2-aminopyridine moiety interacts directly with the catalytic aspartic acids Asp32 and Asp228 via a hydrogen-bonding network. 相似文献
993.
994.
Duane L. Guernsey Haiyan Jiang Karen Bedard Susan C. Evans Meghan Ferguson Makoto Matsuoka Christine Macgillivray Mathew Nightingale Scott Perry Andrea L. Rideout Andrew Orr Mark Ludman David L. Skidmore Timothy Benstead Mark E. Samuels 《PLoS genetics》2010,6(8)
Charcot-Marie-Tooth disease (CMT) represents a family of related sensorimotor neuropathies. We studied a large family from a rural eastern Canadian community, with multiple individuals suffering from a condition clinically most similar to autosomal recessive axonal CMT, or AR-CMT2. Homozygosity mapping with high-density SNP genotyping of six affected individuals from the family excluded 23 known genes for various subtypes of CMT and instead identified a single homozygous region on chromosome 9, at 122,423,730–129,841,977 Mbp, shared identical by state in all six affected individuals. A homozygous pathogenic variant was identified in the gene encoding leucine rich repeat and sterile alpha motif 1 (LRSAM1) by direct DNA sequencing of genes within the region in affected DNA samples. The single nucleotide change mutates an intronic consensus acceptor splicing site from AG to AA. Direct analysis of RNA from patient blood demonstrated aberrant splicing of the affected exon, causing an obligatory frameshift and premature truncation of the protein. Western blotting of immortalized cells from a homozygous patient showed complete absence of detectable protein, consistent with the splice site defect. LRSAM1 plays a role in membrane vesicle fusion during viral maturation and for proper adhesion of neuronal cells in culture. Other ubiquitin ligases play documented roles in neurodegenerative diseases. LRSAM1 is a strong candidate for the causal gene for the genetic disorder in our kindred. 相似文献
995.
Alexandra Pavlova Faith M. Walker Rodney van der Ree Silvana Cesarini Andrea C. Taylor 《Conservation Genetics》2010,11(6):2393-2407
The squirrel glider Petaurus norfolcensis occurs across a broad Australian latitudinal range that includes gaps in distribution and potential biogeographic barriers,
creating the potential for evolution of distinct entities within this species. Because of the species’ threatened status in
the southern part of its range, we tested for the presence of geographically based independent evolutionary units among gliders
sampled from southern, and northern coastal populations, using sequences of mitochondrial cytochrome b DNA (mtDNA) and a set of five nuclear microsatellites in 258 individuals. Our analyses suggest that an initial northward
colonisation in the early- to mid-Pleistocene was followed by isolation by distance and, eventually, divergence between the
sampled coastal and southern populations in the mid- to late-Pleistocene. We propose that the previously large and diverse
southern populations have declined coincidentally with the replacement of wet forests by open sclerophyll woodlands during
the preceding few million years. By contrast coastal populations further north appear to have been expanding and at present
have an effective population size several times greater than southern populations. These results suggest that the two forms
are on different evolutionary trajectories and should be treated separately for conservation purposes. It is highly desirable
that loss of southern populations be prevented to maintain the unique genetic diversity accumulated over a considerable evolutionary
timescale. 相似文献
996.
Proteinase activated receptors (PAR 1-4) are membrane receptors with a unique way of activation by proteinases like thrombin, trypsin and matrix metalloproteinases which lead to a specific cellular response. To evaluate the significance of expression and co-expression of PAR in cancer we performed a survey on published data. A Pubmed literature search on “PAR, thrombin, cancer” was performed and 46 publications were selected for systematic review based on the availability of information on tumor type, material type, detection method and specification of positive cases. PAR-1 was found in 77.3% of malignant samples (n = 678), PAR-2 in 79.5% (n = 592), PAR-3 in 12.6% (n = 87) and PAR-4 in 54.9% (n = 153). PAR-1 and -2 were present in adenocarcinomas, melanomas, osteosarcomas, glioblastomas, meningiomas, leukaemias and squamous cell carcinomas. Presence of PAR-3 was limited to kidney and liver cancer. The data on PAR-4 expression was inconclusive. Those studies analysing PAR-1 and PAR-2 reported coexpression of the two receptors. PAR-1 and -2 are widely expressed in human tumors suggesting an important role in tumorigenesis and providing potential targets for therapy. PAR-3 and PAR-4 are less frequently detectable, their expression and potential role in tumorigenesis require further investigation. 相似文献
997.
Samuel Taddese Michael C. JungChristian Ihling Andrea HeinzReinhard H.H. Neubert Christian E.H. Schmelzer 《Biochimica et Biophysica Acta - Proteins and Proteomics》2010,1804(4):731-739
Collagens of either soft connective or mineralized tissues are subject to continuous remodeling and turnover. Undesired cleavage can be the result of an imbalance between proteases and their inhibitors. Owing to their superhelical structure, collagens are resistant to many proteases and matrix metalloproteinases (MMPs) are required to initiate further degradation by other enzymes. Several MMPs are known to degrade collagens, but the action of MMP-12 has not yet been studied in detail. In this work, the potential of MMP-12 in recognizing sites in human skin collagen types I and III has been investigated. The catalytic domain of MMP-12 binds to the triple helix and cleaves the typical sites -Gly775-Leu776- in α-2 type I collagen and -Gly775-Ile776- in α-1 type I and type III collagens and at multiple other sites in both collagen types. Moreover, it was observed that the region around these typical sites contains comparatively less prolines, of which some have been proven to be only partially hydroxylated. This is of relevance since partial hydroxylation in the vicinity of a potential scissile bond may have a local effect on the conformational thermodynamics with probable consequences on the collagenolysis process. Taken together, the results of the present work confirm that the catalytic domain of MMP-12 alone binds and degrades collagens I and III. 相似文献
998.
Debora Paris Dominique Melck Matteo Stocchero Oceania D’Apolito Rosa Calemma Giuseppe Castello Francesco Izzo Giuseppe Palmieri Gaetano Corso Andrea Motta 《Metabolomics : Official journal of the Metabolomic Society》2010,6(3):405-416
Human hepatocellular carcinoma (HCC) is the most recurrent malignancy of the liver and represents one of the main causes of
cancer death worldwide. Furthermore, the liver is the most frequent site of metastatic colonization, and hepatic metastases
are far more common than primary cancers in Western countries. A possible way of investigating liver diseases is to study
the tissue metabolic profiles. High-resolution nuclear magnetic resonance (NMR) spectroscopy of hepatic tissue extracts was
combined with pattern-recognition and visualization techniques to uncover metabolic differences among analyzed tissue types.
Extracts were from primary HCC, chronic hepatitis-C-virus related cirrhotic tissues, hepatic metastases from colorectal carcinomas,
and non-cirrhotic normal liver tissues adjacent to metastases as controls. We identified all metabolites present in the NMR
spectra, and after statistical evaluation of all spectral classes, we were able to define the metabolic changes underlying
the different liver conditions and diseases. In particular, the lactate and the glucose tissue signals were found to primarily
discriminate the different histological samples. We followed the biochemical changes of human hepatic lesions through primary
(HCC) and secondary (metastases from colorectal carcinoma) liver tumors, cirrhotic tissues, and non-cirrhotic histologically-confirmed
normal liver tissues adjacent to metastases, achieving a metabolic differentiation of the various pathological states based
upon the variation of the intracellular lactate/glucose ratio. It is suggested that such a signal pattern may act as a potential
marker for assessing pathological hepatic lesions. 相似文献
999.
Barnickel B Bayliffe F Diestel R Kempf K Laschat S Pachali S Sasse F Schlenk A Schobert R 《化学与生物多样性》2010,7(12):2830-2845
Fragments and synthetic precursors prepared en route to the macrocyclic 3-acyltetramic acids (=3-acyl-1,5-dihydro-4-hydroxy-2H-pyrrol-2-ones) aburatubolactam and macrocidin A, as well as other analogs with variance in the ring heteroatom (N, O, S), and the residues at N(1), C(3), and C(5) were tested for cytotoxic and antimicrobial effects. Anticancer activity against various tumor cell lines in vitro did not necessarily require an intact pyrrolidin-2,4-dione ring. An acyclic β-hydroxy-octatrienoyl amide precursor to aburatubolactam also exhibited distinct activity with an IC?? (120?h) value of <2.5?μM. The length of 3-oligoenoyl residues had little influence on the anticancer activity, but 3-alka-oligoenoyl tetramic acids were far more efficacious than their 3-(4-methoxycinnamoyl) congeners. N-H-3-acyltetramic acids were generally more active than their N-Me or N-Boc analogs, unless further polar groups necessitated an increased lipophilicity for sufficient uptake. Tetronic and thiotetronic acids were far less antiproliferative in cancer cells when compared with identically substituted tetramic acids. 相似文献
1000.
Federica Susta Davide Chiasserini Katia Fettucciari Pier Luigi Orvietani Flavia Quotadamo Rosina Noce Andrea Bartoli Pierfrancesco Marconi Lanfranco Corazzi Luciano Binaglia 《Proteomics》2010,10(11):2099-2112
Protein expression changes induced in thioglycolate‐elicited peritoneal murine macrophages (MΦ) by infection with type III Group B Streptococcus (GBS) are described. Proteins from control MΦ and MΦ incubated 2 h with live or heat‐inactivated GBS were separated by 2‐DE. Proteins whose expression was significantly different in infected MΦ, as compared with control cells, were identified by MS/MS analysis. Changes in the expression level of proteins involved in both positive and negative modulation of phagocytic functions, stress response and cell death were induced in MΦ by GBS infection. In particular, expression of enzymes playing a key role in production of reactive oxygen species was lowered in GBS‐infected MΦ. Significant alterations in the expression of some metabolic enzymes were also observed, most of the glycolytic and of the pentose‐cycle enzymes being down‐regulated in MΦ infected with live GBS. Finally, evidence was obtained that GBS infection affects the expression of enzymes or enzyme subunits involved in ATP synthesis and in adenine nucleotides interconversion processes. 相似文献