全文获取类型
收费全文 | 2329篇 |
免费 | 174篇 |
国内免费 | 3篇 |
专业分类
2506篇 |
出版年
2022年 | 16篇 |
2021年 | 24篇 |
2020年 | 10篇 |
2019年 | 15篇 |
2018年 | 22篇 |
2017年 | 28篇 |
2016年 | 42篇 |
2015年 | 76篇 |
2014年 | 73篇 |
2013年 | 125篇 |
2012年 | 142篇 |
2011年 | 124篇 |
2010年 | 105篇 |
2009年 | 80篇 |
2008年 | 130篇 |
2007年 | 101篇 |
2006年 | 117篇 |
2005年 | 112篇 |
2004年 | 96篇 |
2003年 | 104篇 |
2002年 | 80篇 |
2001年 | 87篇 |
2000年 | 81篇 |
1999年 | 64篇 |
1998年 | 31篇 |
1997年 | 21篇 |
1996年 | 38篇 |
1995年 | 33篇 |
1994年 | 29篇 |
1993年 | 22篇 |
1992年 | 55篇 |
1991年 | 43篇 |
1990年 | 25篇 |
1989年 | 23篇 |
1988年 | 28篇 |
1987年 | 28篇 |
1986年 | 21篇 |
1985年 | 24篇 |
1984年 | 23篇 |
1983年 | 18篇 |
1982年 | 10篇 |
1981年 | 11篇 |
1979年 | 14篇 |
1978年 | 10篇 |
1977年 | 18篇 |
1976年 | 17篇 |
1974年 | 25篇 |
1973年 | 13篇 |
1972年 | 13篇 |
1970年 | 10篇 |
排序方式: 共有2506条查询结果,搜索用时 16 毫秒
71.
Protein surface charges and Ca2+ binding to individual sites in calbindin D9k: stopped-flow studies 总被引:3,自引:0,他引:3
The kinetics of calcium dissociation from two groups of site-specific mutants of calbindin D9k--a protein in the calmodulin superfamily with two Ca2+ sites and a tertiary structure closely similar to that of the globular domains of troponin C and calmodulin--have been studied by stopped-flow kinetic methods, using the fluorescent calcium chelator Quin 2, and by 43Ca NMR methods. The first group of mutants comprises all possible single, double, and triple neutralizations of three particular carboxylate groups (Glu-17, Asp-19, and Glu-26) that are located on the surface of the protein. These carboxylates are close to the two EF-hand calcium binding sites, but are not directly liganded to the Ca2+ ions. Conservative modification of these negative carboxylate side chains by conversion to the corresponding amides results in a marked reduction in the Ca2+ binding constants for both sites, as recently reported [Linse et al. (1988) Nature 335, 651-652]. The stopped-flow kinetic results show that this reduction in Ca2+ affinity derives primarily from a reduction in the Ca2+ association rate constant, kon. The estimated maximum value of the association rate constant (kon(max) for Ca2+ binding to the wild-type protein is ca. 10(9) M-1 s-1. In contrast, for the mutant protein with three charges neutralized the maximum association rate constant is estimated to be only 2 X 10(7) M-1 s-1.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
72.
The freezing point of aqueous two-phase (liquid-liquid) systems containing water, dextran and poly(ethylene glycol) has been lowered by including glycerol. Biological membranes, obtained by fragmentation of a crude synaptosomal preparation from calf brain cortex, have been included in the two-phase systems. The effects of temperature and the concentration of glycerol on the partition of the membranes within the systems have been investigated. Considerable stabilisation of the membranes was noticed when they were partitioned at -10 degrees C compared with 0 degrees C. The influences of glycerol, ethylene glycol, N,N-dimethylformamide and tetrahydrofuran on the phase-forming properties of the systems and on enzyme activities are also presented. Possible use of the above systems for studies and separation of biological membranes are discussed. 相似文献
73.
P Brodin C Johansson S Forsén T Drakenberg T Grundstr?m 《The Journal of biological chemistry》1990,265(19):11125-11130
We have constructed three different engineered proteins based on calbindin D9K by either exchanging the two calcium binding sites within the protein or making the amino acid sequence of the two calcium binding sites identical. The individual calcium binding sites of the engineered proteins retain most of their ion binding characteristics as well as the basal structure of their Ca2+ ligand sphere in the new environment. Even the protein with its sites interchanged, a mutation involving 30 amino acids out of a total of 75, still binds calcium with an affinity as high as that of many natural EF-hand proteins. 相似文献
74.
We have studied the changes in the activities of both nitrogenase (switch off) and glutamine synthetase in Rhodospirillum rubrum upon addition of ammonium ions or glutamine to nitrogen fixing cultures. Both activities decrease drastically and return in a parallel manner when added ammonia is metabolized. The decrease in glutamine synthetase activity does not seem to be primarily due to adenylylation of the enzyme. Addition of glutamine to cells starved for nitrogen results in inactivation of glutamine synthetase but nitrogenase is only partially switched off.Abbreviations CeMe3NBr
Cetyltrimethylammonium bromide
- Hepes
N-2-hydroxyethyl-piperazine-N-2 sulfonic acid
- MSO
methionine-D,L-sulfoximine
- Tea-Dmg
triethanol amine-3,3-dimethylglutaric acid 相似文献
75.
Studies of chromophores in model membranes by polarized light-absorption spectroscopy. The orientation and binding of tetracaine and procaine.
下载免费PDF全文
![点击此处可从《Biophysical journal》网站下载免费的PDF全文](/ch/ext_images/free.gif)
A method for studying the orientation and binding of chromophores in macroscopically aligned membranes by polarized light absorption spectroscopy is described. Here tetracaine and procaine solubilized in a lamellar phase of octanoyl-1-glyceride (monooctanoin) and water have been investigated. Tetracaine is found to be located in the lipid region with a preferential orientation of the molecular long axis parallel to the hydrocarbon chains. The orientation of procaine, mainly residing in the water region, is very small. 相似文献
76.
A covalently bound photoisomerizable agonist. Comparison with reversibly bound agonists at electrophorus electroplaques 总被引:2,自引:2,他引:0
下载免费PDF全文
![点击此处可从《The Journal of general physiology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
HA Lester ME Krouse MM Nass NH Wassermann BF Erlanger 《The Journal of general physiology》1980,75(2):207-232
After disulphide bonds are reduced with dithiothreitol, trans-3- (α-bromomethyl)-3’-[α- (trimethylammonium)methyl]azobenzene (trans-QBr) alkylates a sulfhydryl group on receptors. The membrane conductance induced by this “tethered agonist” shares many properties with that induced by reversible agonists. Equilibrium conductance increases as the membrane potential is made more negative; the voltage sensitivity resembles that seen with 50 [mu]M carbachol. Voltage- jump relaxations follow an exponential time-course; the rate constants are about twice as large as those seen with 50 μM carbachol and have the same voltage and temperature sensitivity. With reversible agonists, the rate of channel opening increases with the frequency of agonist-receptor collisions: with tethered trans-Qbr, this rate depends only on intramolecular events. In comparison to the conductance induced by reversible agonists, the QBr-induced conductance is at least 10-fold less sensitive to competitive blockade by tubocurarine and roughly as sensitive to “open-channel blockade” bu QX-222. Light-flash experiments with tethered QBr resemble those with the reversible photoisomerizable agonist, 3,3’,bis-[α-(trimethylammonium)methyl]azobenzene (Bis-Q): the conductance is increased by cis {arrow} trans photoisomerizations and decreased by trans {arrow} cis photoisomerizations. As with Bis-Q, ligh-flash relaxations have the same rate constant as voltage-jump relaxations. Receptors with tethered trans isomer. By comparing the agonist-induced conductance with the cis/tans ratio, we conclude that each channel’s activation is determined by the configuration of a single tethered QBr molecule. The QBr-induced conductance shows slow decreases (time constant, several hundred milliseconds), which can be partially reversed by flashes. The similarities suggest that the same rate-limiting step governs the opening and closing of channels for both reversible and tethered agonists. Therefore, this step is probably not the initial encounter between agonist and receptor molecules. 相似文献
77.
The interaction of human serum albumin with fatty acids has been determined using the method of affinity partitioning in aqueous biphasic systems containing dextran, poly(ethylene glycol) and esters of dicarboxylic acids with poly(ethylene glycol). The difference in the partition of albumin in phase systems with and without the poly(ethylene glycol)-bound fatty acid group provides a measure of the interaction of fatty acids with the protein. The relative contribution of the polar and non-polar interaction to the binding of fatty acids to albumin has been estimated by comparing the present data with that obtained earlier using poly(ethylene glycol)-bound straight chain aliphatic hydrocarbons. In both cases, the aliphatic chain should contain a minimum of 8 carbon atoms to affect the partition of albumin and that the maximum effect is obtained with chains containing 16 carbon atoms. The effect of the polymer-bound fatty acid group is higher than the corresponding hydrocarbon only when the number of carbon atoms in it exceeds 12. The relative effect of polymer-bound 16-carbon chains, with and without a carboxyl group in the terminal position is independent of pH in the range 5--9. 相似文献
78.
An aqueous polymer phase system containing 6.3% (w/w) dextran and 3.5% (w/w) poly(ethylene glycol) in 10 mM phosphate buffer (pH 8.0) was developed to select RNA-DNA hybrids from unhybridized RNA. The top phase of this phase system, which contains DNA and the RNA-DNA hybrids, can be used to purify adenovirus messenger RNA both early and late in the infectious cycle. The hybrids can be melted by heat in the top phase and the messenger RNA selected by oligo(dT)cellulose chromatography whereupon the polymers and the DNA percolate and the polyadenylated messenger RNA absorb to the column. The isolated messenger RNA appears to be almost quantitatively recovered at a purity from 70 to 90% depending on the concentration of the specific messenger RNA in the starting material. Early and late viral messenger RNA were selected on the complementary strands of adenovirus DNA according to this procedure. 相似文献
79.
The surface properties of spinach chloroplasts, both of intact chloroplasts with surrounding envelope and broken chloroplasts consisting of the inner lamellar system, have been studied by partitioning them between two aqueous phases, especially using counter-current distribution technique. The two-phase system consists of poly(ethyleneglycol), dextran and water. The two polymers are enriched in opposite phases and by binding deoxycholate or palmitate to one of the polymers the affinity of chloroplasts for the corresponding phase is strongly enhanced. The partition of the two classes of chloroplasts, however, is not affected to the same degree and the affinity of the chloroplast envelope for deoxycholate and palmitate is stronger than that of the lamellar system. This has been correlated to the chemical composition of the two types of membranes. By studying the effect of salts on the partition it has been found that the lamellar system bears a larger number of negative charges as compared to the envelope of the intact chloroplast. 相似文献
80.
Four of the membrane proteins from Acholeplasma laidlawii that are soluble in the nonionic detergent Tween 20 have been purified by preparative electrophoretic techniques utilizing different supporting media. The last purification step for two of the major proteins was a preparative polyacrylamide gel electrophoresis performed in the absence of any detergent. The proteins were recovered by continuous elution. The purity of the fractions was examined by analytical polyacrylamide gel electrophoresis and crossed immunoelectrophoresis. Two of the minor proteins were purified by dextran gel electrophoresis as the final step, which was also performed in a detergent-free buffer. The separation was followed by scanning the dextran gel in ultraviolet light. The proteins were recovered by slicing the gel and degrading the gel slices with dextranase. The homogeneity of the fractions was checked by electroimmunoassay. 相似文献