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11.
Circular dichroism spectroscopy has been used to investigate the binding of valinomycin to bacteriorhodopsin in purple membrane suspensions. Addition of valinomycin to purple membrane suspensions obtained from Halobacterium halobium causes the circular dichroism spectrum to shift from an aggregate spectrum to one resembling a monomer spectrum, indicating a loss of chromophore-chromophore interactions. By observing the spectral change upon titration of valinomycin, an apparent dissociation constant of 30–40 M for valinomycin binding was determined. Kinetics of dark adaptation for valinomycin-treated purple membrane are comparable to those for monomeric bacteriorhodopsin. Centrifugation studies demonstrate that valinomycin-treated purple membrane sediments the same as untreated purple membrane suspensions. These results are consistent with a model in which valinomycin binds specifically to bacteriorhodopsin without disrupting the purple membrane fragments.Abbreviations BR
bacteriorhodopsin
- CD
circular dichroism
- Tricine
N-[tris-(hydroxymethyl) methyl] glycine 相似文献
12.
Characterization of Gypsy Moth (Lymantria dispar) Nuclear Polyhedrosis Virus 总被引:1,自引:0,他引:1 下载免费PDF全文
Characterization of the proteins and nucleic acid of the gypsy moth nuclear polyhedrosis virus isolated in Ithaca, N.Y. (LdNPV-IT) is presented. A total of 29 viral structural proteins were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis when the virus was isolated in the absence of alkaline protease activity. Fourteen surface envelope viral proteins were identified by lactoperoxidase iodination. Eleven proteins were associated with nucleocapsids prepared by Nonidet P-40 detergent treatment. Distinct alterations of viral proteins were documented when virions were purified in the presence of occlusion body-associated alkaline protease(s). Restriction enzyme digests of viral DNA indicated that this isolate was composed of a large number of genetic variants. On the basis of the major molar fragments resulting from EcoRI, BamHI, BglII, and HindIII digests, the molecular weight of the LdNPV genome was approximately 88 × 106. 相似文献
13.
F S Nallaseth R P Lawther M R Stallcup M J Dewey 《Molecular & general genetics : MGG》1983,190(1):80-84
The mammalian Y chromosome is an isolated piece of genetic material that directs sexual determination and gametogenesis. Very little is understood about the mechanism whereby the Y chromosome carries out these functions. Also, there is a severe lack of genetic markers on this chromosome. In order to understand the structure and function of the Y chromosome at the level of its DNA sequences and to provide genetic markers, we are isolating clones of DNA whose sequences are found primarily in DNA from male mice. To this end, we have developed a procedure for the identification of such clones. Application of this screening procedure to a lambda library derived from mouse sperm DNA has yielded 12 distinct clones, part of whose sequences are present predominantly in male DNA. Besides this DNA, they also contain other sequences that are shared with female DNA. These clones are either derived from the Y chromosome or they represent autosomal sequences specifically amplified during male development. 相似文献
14.
Otto D. Hensens Ray S. Dewey Jerrold M. Liesch Mary A. Napier Robert A. Reamer Jack L. Smith Georg Albers-Schönberg Irving H. Goldberg 《Biochemical and biophysical research communications》1983,113(2):538-547
Spectroscopic evidence suggests the presence of a highly strained ether ring (Fig. 1) (possibly an epoxide) in the C12-subunit of the previously determined partial structure (Fig. 2) of the major neocarzinostatin chromophore (NCS-Chrom ) which completes assignment of all the oxygens in the molecule. The main product from mercaptan treatment suggests opening of the ether ring involving the addition of one molecule of mercaptan as well as reduction of the C12-substructure, whereas a parallel two-step reduction occurs on NaBH4 treatment. Both reactions occur with rearrangement of the C12-substructure and the implication for the mechanism of action of NCS-Chrom in DNA strand scission activity is discussed. The evidence suggests a downward revision of the molecular formula for NCS-Chrom as well as minor components and by two protons. 相似文献
15.
Characterization of soybean choline kinase cDNAs and their expression in yeast and Escherichia coli. 下载免费PDF全文
An expressed sequence tag from Arabidopsis that displayed sequence homology to mammalian and yeast choline kinases was used to isolate choline kinase-like cDNAs from soybean (Glycine max L.). Two distinct cDNAs, designated GmCK1 and GmCK2, were recovered that possessed full-length reading frames, each sharing approximately 32% identity at the predicted amino acid level with the rat choline kinase. A third unique choline kinase-like cDNA, GmCK3, was also identified but was not full length. Heterologous expression of GmCK1 in yeast (Saccharomyces cerevisiae) and GmCK2 in both yeast and Escherichia coli demonstrated that each encodes choline kinase activity. In addition to choline, other potential substrates for the choline kinase enzyme include ethanolamine, monomethylethanolamine (MME), and dimethylethanolamine (DME). Both soybean choline kinase isoforms demonstrated negligible ethanolamine kinase activity. Competitive inhibition assays, however, revealed very distinct differences in their responses to DME and MME. DME effectively inhibited only the GmCK2-encoded choline kinase activity. Although MME failed to effectively inhibit either reaction, an unexpected enhancement of choline kinase activity was observed specifically with the GmCK1-encoded enzyme. These results show that choline kinase is encoded by a small, multigene family in soybean comprising two or more distinct isoforms that exhibit both similarities and differences with regard to substrate specificity. 相似文献
16.
Theoretical estimates of mechanical properties of the endothelial cell cytoskeleton. 总被引:3,自引:0,他引:3 下载免费PDF全文
Current modeling of endothelial cell mechanics does not account for the network of F-actin that permeates the cytoplasm. This network, the distributed cytoplasmic structural actin (DCSA), extends from apical to basal membranes, with frequent attachments. Stress fibers are intercalated within the network, with similar frequent attachments. The microscopic structure of the DCSA resembles a foam, so that the mechanical properties can be estimated with analogy to these well-studied systems. The moduli of shear and elastic deformations are estimated to be on the order of 10(5) dynes/cm2. This prediction agrees with experimental measurements of the properties of cytoplasm and endothelial cells reported elsewhere. Stress fibers can potentially increase the modulus by a factor of 2-10, depending on whether they act in series or parallel to the network in transmitting surface forces. The deformations produced by physiological flow fields are of insufficient magnitude to disrupt cell-to-cell or DCSA cross-linkages. The questions raised by this paradox, and the ramifications of implicating the previously unreported DCSA as the primary force transmission element are discussed. 相似文献
17.
A comprehensive data base is analyzed to determine the Shannon information content of a protein sequence. This information entropy is estimated by three methods: a k-tuplet analysis, a generalized Zipf analysis, and a "Chou-Fasman gambler." The k-tuplet analysis is a "letter" analysis, based on conditional sequence probabilities. The generalized Zipf analysis demonstrates the statistical linguistic qualities of protein sequences and uses the "word" frequency to determine the Shannon entropy. The Zipf analysis and k-tuplet analysis give Shannon entropies of approximately 2.5 bits/amino acid. This entropy is much smaller than the value of 4.18 bits/amino acid obtained from the nonuniform composition of amino acids in proteins. The "Chou-Fasman" gambler is an algorithm based on the Chou-Fasman rules for protein structure. It uses both sequence and secondary structure information to guess at the number of possible amino acids that could appropriately substitute into a sequence. As in the case for the English language, the gambler algorithm gives significantly lower entropies than the k-tuplet analysis. Using these entropies, the number of most probable protein sequences can be calculated. The number of most probable protein sequences is much less than the number of possible sequences but is still much larger than the number of sequences thought to have existed throughout evolution. Implications of these results for mutagenesis experiments are discussed. 相似文献
18.
Allyson Tucker Dilks Joanne Gilchrist Yik Lam Nathan Nicholes Brad Stanley 《Biotechnology progress》2023,39(4):e3351
No-salt flowthrough hydrophobic interaction chromatography (HIC) has been shown to effectively remove process and product-related impurities from bioprocess streams. In this publication, a panel of six antibodies has been used to demonstrate operating principles for the application of no-salt flowthrough HIC in antibody purification processes. The results indicate that no-salt flowthrough HIC provides robust aggregate clearance across operating conditions including flow rate, and variations in resin ligand density. Additionally, HMW reduction has an optimal pH range relative to the isoelectric point of each molecule and high molecular weight (HMW) reduction can be improved by altering the total protein load and/or HMW concentration to drive binding of high molecular weight species to the resin. 相似文献
19.
The Lipid Peroxidation Product, 4-Hydroxy-2-trans-Nonenal, Alters the Conformation of Cortical Synaptosomal Membrane Proteins 总被引:6,自引:1,他引:5
‡Ram Subramaniam ‡Fred Roediger Brad Jordan †‡§Mark P. Mattson §Jeffrey N. Keller Georg Waeg ‡§ D. Allan Butterfield 《Journal of neurochemistry》1997,69(3):1161-1169
Abstract: Alzheimer's disease (AD) is widely held to be a disorder associated with oxidative stress due, in part, to the membrane action of amyloid β-peptide (Aβ). Aβ-associated free radicals cause lipid peroxidation, a major product of which is 4-hydroxy-2- trans -nonenal (HNE). We determined whether HNE would alter the conformation of synaptosomal membrane proteins, which might be related to the known neurotoxicity of Aβ and HNE. Electron paramagnetic resonance spectroscopy, using a protein-specific spin label, MAL-6(2,2,6,6-tetramethyl-4-maleimidopiperidin-1-oxyl), was used to probe conformational changes in gerbil cortical synaptosomal membrane proteins, and a lipid-specific stearic acid label, 5-nitroxide stearate, was used to probe for HNE-induced alterations in the fluidity of the bilayer domain of these membranes. Synaptosomal membranes, incubated with low concentrations of HNE, exhibited changes in protein conformation and bilayer order and motion (fluidity). The changes in protein conformation were found to be concentration- and time-dependent. Significant protein conformational changes were observed at physiologically relevant concentrations of 1–10 µ M HNE, reminiscent of similar changes in synaptosomal membrane proteins from senile plaque- and Aβ-rich AD hippocampal and inferior parietal brain regions. HNE-induced modifications in the physical state of gerbil synaptosomal membrane proteins were prevented completely by using excess glutathione ethyl ester, known to protect neurons from HNE-caused neurotoxicity. Membrane fluidity was found to increase at higher concentrations of HNE (50 µ M ). The results obtained are discussed with relevance to the hypothesis of Aβ-induced free radical-mediated lipid peroxidation, leading to subsequent HNE-induced alterations in the structure and function of key membrane proteins with consequent neurotoxicity in AD brain. 相似文献
20.
Interpreting photoactivated fluorescence microscopy measurements of steady-state actin dynamics. 总被引:1,自引:0,他引:1 下载免费PDF全文
A continuum model describing the steady-state actin dynamics of the cytoskeleton of living cells has been developed to aid in the interpretation of photoactivated fluorescence experiments. In a simplified cell geometry, the model assumes uniform concentrations of cytosolic and cytoskeletal actin throughout the cell and no net growth of either pool. The spatiotemporal evolution of the fluorescent actin population is described by a system of two coupled linear partial-differential equations. An analytical solution is found using a Fourier-Laplace transform and important limiting cases relevant to the design of experiments are discussed. The results demonstrate that, despite being a complex function of the parameters, the fluorescence decay in photoactivated fluorescence experiments has a biphasic behavior featuring a short-term decay controlled by monomer diffusion and a long-term decay governed by the monomer exchange rate between the polymerized and unpolymerized actin pools. This biphasic behavior suggests a convenient mechanism for extracting the parameters governing the fluorescence decay from data records. These parameters include the actin monomer diffusion coefficient, filament turnover rate, and ratio of polymerized to unpolymerized actin. 相似文献