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171.
Hölzl H Kapelari B Kellermann J Seemüller E Sümegi M Udvardy A Medalia O Sperling J Müller SA Engel A Baumeister W 《The Journal of cell biology》2000,150(1):119-130
Drosophila melanogaster embryos are a source for homogeneous and stable 26S proteasomes suitable for structural studies. For biochemical characterization, purified 26S proteasomes were resolved by two-dimensional (2D) gel electrophoresis and subunits composing the regulatory complex (RC) were identified by amino acid sequencing and immunoblotting, before corresponding cDNAs were sequenced. 17 subunits from Drosophila RCs were found to have homologues in the yeast and human RCs. An additional subunit, p37A, not yet described in RCs of other organisms, is a member of the ubiquitin COOH-terminal hydrolase family (UCH). Analysis of EM images of 26S proteasomes-UCH-inhibitor complexes allowed for the first time to localize one of the RC's specific functions, deubiquitylating activity.The masses of 26S proteasomes with either one or two attached RCs were determined by scanning transmission EM (STEM), yielding a mass of 894 kD for a single RC. This value is in good agreement with the summed masses of the 18 identified RC subunits (932 kD), indicating that the number of subunits is complete. 相似文献
172.
173.
Summary We report on some aspects of the breeding biology of the critically endangered Writhed-billed Hornbill (Aceros waldeni) on the island of Panay, Philippines. Observations were made at three nests during 1995–1997. Walling-in of the females commenced in the first week of March. One female remained incarcerated for 77 days, two of three broods completed fledging around May 20 (1995, 1997). Details on fledging of the female and her brood and postfledging care by both parents are reported.The food of the males at two nests was ca. 98% fruits and 2% invertebrates. The plants exploited comprised at least 14 species. Over a third of the fruits delivered were figs of a small number of species.Two males had average feeding rates of 0.56 and 0.88 times per hour respectively, and fed 1 to 66 (median 8) items per feeding visit at the nest. The hourly feeding rate increased after hatching, but the composition of the diet did not change noticeably. As a rule, food items were delivered singly and, during one visit, in runs of one, or rarely up to 3, species.In the three weeks following vacation of the nest, the male appeared to be the sole food provider while the female stayed continually with the 3 young (as sentinel?) in the vicinity of the nest.The nest environs were defended by the male against Tarictic Hornbills (Penelopides panini panini). Six vocalisations of the parents are mentioned. One was used in territorial skirmishes with Tarictic Hornbills.With perhaps less than 30 pairs of the Writhed-bill surviving, the future for the species looks bleak. Only drastic conservation measures can prevent the species' demise. Some have been started by the PESCP.This paper is publication No. 11 of the Philippine Endemic Species Conservation Project (PESCP) of the Frankfurt Zoological Society. 相似文献
174.
Mayr J Seemüller E Müller SA Engel A Baumeister W 《Journal of structural biology》1998,124(2-3):179-188
Electron microscopy and STEM mass measurements have been used to characterize late intermediates in the assembly pathway of wildtype and mutant Rhodococcus proteasomes. A proteolytically inactive and processing-incompetent mutant, betaK33A, allowed a short-lived late intermediate of the pathway to be captured, the preholoproteasome. In this fully assembled 20S complex the 14 propeptides with an aggregate mass of 100 kDa fill the whole central cavity and most of the two antechambers. It is further shown that in wildtype Rhodococcus proteasomes the propeptides are degraded in a processive manner undergoing multiple cleavages before the products are discharged and the inner cavities are cleared. It appears that the docking of two half-proteasomes, i.e., preholoproteasome formation, is sufficient to trigger autocleavage of the Gly-1/Thr1 bond necessary for active site formation and the subsequent degradation of the propeptides. 相似文献
175.
All-trans retinol, vitamin D and other hydrophobic compounds bind in the axial pore of the five-stranded coiled-coil domain of cartilage oligomeric matrix protein. 总被引:1,自引:0,他引:1
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Y Guo D Bozic V N Malashkevich R A Kammerer T Schulthess J Engel 《The EMBO journal》1998,17(18):5265-5272
The potential storage and delivery function of cartilage oligomeric matrix protein (COMP) for cell signaling molecules was explored by binding hydrophobic compounds to the recombinant five-stranded coiled-coil domain of COMP. Complex formation with benzene, cyclohexane, vitamin D3 and elaidic acid was demonstrated through increases in denaturation temperatures of 2-10 degreesC. For all-trans retinol and all-trans retinoic acid, an equilibrium dissociation constant KD = 0.6 microM was evaluated by fluorescence titration. Binding of benzene and all-trans retinol into the hydrophobic axial pore of the COMP coiled-coil domain was proven by the X-ray crystal structures of the corresponding complexes at 0.25 and 0.27 nm resolution, respectively. Benzene binds with its plane perpendicular to the pore axis. The binding site is between the two internal rings formed by Leu37 and Thr40 pointing into the pore of the COMP coiled-coil domain. The retinol beta-ionone ring is positioned in a hydrophobic environment near Thr40, and the 1.1 nm long isoprene tail follows a completely hydrophobic region of the pore. Its terminal hydroxyl group complexes with a ring of the five side chains of Gln54. A mutant in which Gln54 is replaced by Ile binds all-trans retinol with affinity similar to the wild-type, demonstrating that hydrophobic interactions are predominant. 相似文献
176.
Mac-2 binding protein is a cell-adhesive protein of the extracellular matrix which self-assembles into ring-like structures and binds beta1 integrins, collagens and fibronectin. 总被引:1,自引:0,他引:1
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Human Mac-2 binding protein (M2BP) was prepared in recombinant form from the culture medium of 293 kidney cells and consisted of a 92 kDa subunit. The protein was obtained in a native state as indicated by CD spectroscopy, demonstrating alpha-helical and beta-type structure, and by protease resistance and immunological analysis. It was highly modified by N- and O-glycosylation but not by glycosaminoglycans. Ultracentrifugation showed non-covalent association into oligomers with molar masses of 1000-1500 kDa. Electron microscopy showed ring-like shapes with diameters of 30-40 nm. M2BP bound in solid-phase assays to collagens IV, V and VI, fibronectin and nidogen, but not to fibrillar collagens I and III or other basement membrane proteins. The protein also mediated adhesion of cell lines at comparable strength with laminin. Adhesion to M2BP was inhibited by antibodies to integrin beta1 subunits but not to alpha2 and alpha6 subunits, RGD peptide or lactose. This distinguishes cell adhesion of M2BP from that of laminin and excludes involvement of lactose-binding galectin-3. Immunological assays demonstrated variable secretion by cultured human cells of M2BP, which was detected in the extracellular matrix of several mouse tissues. 相似文献
177.
178.
Willem H. van de Poll Engel G. Vrieling Winfried W. C. Gieskes 《Journal of phycology》1999,35(5):1044-1053
A detailed immunocytochemical and biochemical study of the location and expression of frustulins, a family of proteins associated with the frustules of diatoms, has been performed for Cylindrotheca fusiformis Reimann et Lewin, Navicula pelliculosa (Brébisson et Kützing) Hilse, and Navicula salinarum (Grunow) Husted. Immunocytochemistry revealed that frustulins, which share homologous epitopes but are different in size, were predominantly located in the organic casing. Based on timed immunolocalization experiments and Western blotting analysis of cell extracts obtained sequentially after repleting silicate to Si-synchronized cells, the continuous presence of the frustulins in the mature and parental organic casing of the examined species was observed. The frustulins of N. pelliculosa appeared as proteins similar to those of C. fusiformis, sharing identical epitopes. The extractions, however, yielded a markedly lower abundance of frustulins in N. pelliculosa. Peak concentrations of extracted frustulins appeared to be expressed just ahead of the silicification process in C. fusiformis, whereas the level of expression in N. pelliculosa increased along with maturation of the new valves. For N. salinarum, the presence of the frustulins could not be confirmed properly by Western blotting, most probably because of the small sample volumes, inefficient extraction, and a lower amount of homologous frustulins in the casing of this species. It is concluded that the frustulins of these species are not associated with the silicalemma of the newly formed silica deposition vesicles and therefore do not seem to be involved in the silicification process itself. Overall, the results imply a structural role of the frustulins in the casing of diatoms rather than a regulation of the silicification process. 相似文献
179.
Natacha Roudnitzky Maik Behrens Anika Engel Susann Kohl Sophie Thalmann Sandra Hübner Kristina Lossow Stephen P. Wooding Wolfgang Meyerhof 《PLoS genetics》2015,11(9)
The ability to taste bitterness evolved to safeguard most animals, including humans, against potentially toxic substances, thereby leading to food rejection. Nonetheless, bitter perception is subject to individual variations due to the presence of genetic functional polymorphisms in bitter taste receptor (TAS2R) genes, such as the long-known association between genetic polymorphisms in TAS2R38 and bitter taste perception of phenylthiocarbamide. Yet, due to overlaps in specificities across receptors, such associations with a single TAS2R locus are uncommon. Therefore, to investigate more complex associations, we examined taste responses to six structurally diverse compounds (absinthin, amarogentin, cascarillin, grosheimin, quassin, and quinine) in a sample of the Caucasian population. By sequencing all bitter receptor loci, inferring long-range haplotypes, mapping their effects on phenotype variation, and characterizing functionally causal allelic variants, we deciphered at the molecular level how a subjects’ genotype for the whole-family of TAS2R genes shapes variation in bitter taste perception. Within each haplotype block implicated in phenotypic variation, we provided evidence for at least one locus harboring functional polymorphic alleles, e.g. one locus for sensitivity to amarogentin, one of the most bitter natural compounds known, and two loci for sensitivity to grosheimin, one of the bitter compounds of artichoke. Our analyses revealed also, besides simple associations, complex associations of bitterness sensitivity across TAS2R loci. Indeed, even if several putative loci harbored both high- and low-sensitivity alleles, phenotypic variation depended on linkage between these alleles. When sensitive alleles for bitter compounds were maintained in the same linkage phase, genetically driven perceptual differences were obvious, e.g. for grosheimin. On the contrary, when sensitive alleles were in opposite phase, only weak genotype-phenotype associations were seen, e.g. for absinthin, the bitter principle of the beverage absinth. These findings illustrate the extent to which genetic influences on taste are complex, yet arise from both receptor activation patterns and linkage structure among receptor genes. 相似文献
180.