全文获取类型
收费全文 | 1345篇 |
免费 | 139篇 |
出版年
2022年 | 7篇 |
2021年 | 16篇 |
2020年 | 11篇 |
2019年 | 12篇 |
2018年 | 16篇 |
2017年 | 17篇 |
2016年 | 33篇 |
2015年 | 44篇 |
2014年 | 61篇 |
2013年 | 58篇 |
2012年 | 82篇 |
2011年 | 77篇 |
2010年 | 59篇 |
2009年 | 45篇 |
2008年 | 70篇 |
2007年 | 75篇 |
2006年 | 67篇 |
2005年 | 79篇 |
2004年 | 84篇 |
2003年 | 57篇 |
2002年 | 56篇 |
2001年 | 48篇 |
2000年 | 64篇 |
1999年 | 44篇 |
1998年 | 25篇 |
1997年 | 14篇 |
1996年 | 14篇 |
1995年 | 11篇 |
1994年 | 15篇 |
1993年 | 7篇 |
1992年 | 28篇 |
1991年 | 28篇 |
1990年 | 22篇 |
1989年 | 22篇 |
1988年 | 18篇 |
1987年 | 18篇 |
1986年 | 12篇 |
1985年 | 4篇 |
1984年 | 4篇 |
1983年 | 3篇 |
1981年 | 6篇 |
1980年 | 6篇 |
1979年 | 6篇 |
1978年 | 7篇 |
1977年 | 5篇 |
1976年 | 7篇 |
1975年 | 3篇 |
1968年 | 2篇 |
1965年 | 3篇 |
1959年 | 2篇 |
排序方式: 共有1484条查询结果,搜索用时 15 毫秒
21.
The mechanism by which cAMP modulates the activity of phosphoinositide-specific phospholipase C (PLC) was studied. Elevation of cAMP inhibited both basal and norepinephrine-stimulated phosphoinositide breakdown in C6Bu1 cells which contain at least three PLC isozymes, PLC-beta, PLC-gamma, and PLC-delta. Treatment of C6Bu1 cells with cAMP-elevating agents (cholera toxin, isobutylmethylxanthine, forskolin, and 8-bromo-cAMP) increased serine phosphate in PLC-gamma, but the phosphate contents in PLC-beta and PLC-delta were not changed. In addition, cAMP-dependent protein kinase selectively phosphorylated purified PLC-gamma among the three isozymes and added a single phosphate at serine. The serine phosphorylation, nevertheless, did not affect the activity of PLC-gamma in vitro. We propose, therefore, that the phosphorylation of PLC-gamma by cAMP-dependent protein kinase alters its interaction with putative modulatory proteins and leads to its inhibition. 相似文献
22.
Tyrosine phosphorylation of phospholipase C-II in vitro by the epidermal growth factor receptor 总被引:23,自引:0,他引:23
S Nishibe M I Wahl S G Rhee G Carpenter 《The Journal of biological chemistry》1989,264(18):10335-10338
In a number of cell lines, epidermal growth factor (EGF) rapidly stimulates the breakdown of inositol phospholipids. Phosphatidylinositol-specific phospholipase C (PLC), therefore, plays an important role in this biological response to EGF, but the mechanism by which EGF-receptor complexes modulate the activation of PLC is not understood. We have previously suggested that tyrosine phosphorylation of PLC or an unknown PLC-associated protein by the EGF receptor is involved in the activation process (Wahl, M. I., Daniel, T. O., and Carpenter, G. (1988) Science 241, 968-970) and have recently shown by immunoprecipitation that the addition of EGF to 32P-labeled cells increases tyrosine and serine phosphorylation of PLC-II (Wahl, M. I., Nishibe, S., Suh, P.-G., Rhee, S. G., and Carpenter, G. (1989) Proc. Natl. Acad. Sci. U.S.A. 86, 1568-1572). In this communication we demonstrate that PLC-II (Mr = 145,000) purified from bovine brain can be phosphorylated in vitro in an EGF-dependent manner by the tyrosine kinase activity of the purified EGF receptor. While PLC-II is an efficient phosphorylation substrate for the purified EGF receptor, PLC-I is a poor substrate and PLC-III is not phosphorylated to any detectable extent. Though all three PLC isozymes possess typical tyrosine phosphorylation sequences, the EGF receptor is surprisingly selective in vitro for the phosphorylation of PLC-II. High performance liquid chromatography comparison of tryptic phosphotyrosyl peptides from PLC-II phosphorylated in vivo and in vitro indicated a similar pattern of multiple tyrosine phosphorylation sites. These findings show that the EGF receptor can directly phosphorylate PLC-II in an efficient and selective manner. 相似文献
23.
Summary A heterologous gene from bloodsucking leech for anticoagulant, hirudin, has been expressed in the methylotrophic yeast Hansenula polymorpha. The addition of 1%(v/v) soybean oil to the medium as a stabilizer enhanced the expression of the hirudin gene in H. polymorpha with MOX promoter. The production of hirudin in the medium with soybean oil was 320mg/L in a 5L fermenter. 相似文献
24.
Summary A genetically modified levansucrase, which contained His-affinity tag in its C-terminal, was constructed by PCR reaction using two synthetic primers. This modified protein was produced up to 30 % in total cell protein of E. coli, and was purified by a one-step affinity chromatography. The optimum pH for levan production was pH 5 and the optimum temperature was 0 °C. The higher velocity of levan formation within shorter enzyme reaction times was achieved by increasing the levels of enzyme concentration. The optimal sucrose concentration for levan production was around 20 %. Under these conditions, more than 50 g levan/l was produced. 相似文献
25.
Batch and continuous hydrolysis of olive oil in an organic-aqueous two-phase system using the live whole cell of Pseudomonas putida 3SK as a source of a lipase is investigated. The strain was not only fully viable and grown well, but also produced extracellular lipase simultaneously. The degree of hydrolysis, depending on olive oil concentration in the solvents, was maximal at 13.5% (w/v) and decreased with the increase of the substrate concentration. At the optimal condition, a degree of hydrolysis higher than 95% was achieved with 24 h at 30 degrees C when the reaction was carried out in a two-phase batch stirred reactor. For long-term operation a continuous stirred reactor was designed. When the reaction was carried out in a continuous stirred reactor, the degree was hydrolysis reached 86% at a dilution rate of 0.2 h(-1). Satisfactory performance of a two-phase bioreactor was obtained in a long-term continous operation, which lasted for at least 30 days by feeding organic solvent containing olive oil and aqueous media separately. (c) 1994 John Wiley & Sons, Inc. 相似文献
26.
27.
SPA::EcoRI fusion protein was produced by Escherichia coli JM103 carrying the multicopy expression plasmid pMTC48, the multicopy repressor plasmid pRK248, and the multicopy protection plasmid pEcoR4 in a 60-L working volume airlift tower loop reactor on M9 minimal medium with glucose. Cell mass concentration, total cell count, number of colony-forming units, specific growth rate, yield coefficient, and metabolite (acetate, pyruvate, succinate, lactate, ethanol) concentrations were monitored during the growth phase and gene expression. Gene expression was induced by temperature shift or chemically by isopropyl-thiogalactosidase in the airlift tower loop reactor (ALTR) at constant cultivation time and in a small stirred tank reactor at different cultivation times. During induction, the cultivation medium was supplemented with concentrated Luria-Bertani (LB) medium. The intracellular enzyme activity was evaluated as a function of the time after the start of the induction. It was found that the reduction of the glucose concentration and increase of the dissolved oxygen concentration reduced the acetate produced and increased the intracellular enzyme activity. (c) 1993 John Wiley & Sons, Inc. 相似文献
28.
A bacterium which has phospholipase A1 activity was isolated from soil. It is aerobic, motile, oxidase-negative and has flagella. The G+C content of the DNA was 58.1 mol%. The major isoprenoids of cell was were Q-8 and MK-8. The main cellular fatty acids were saturated straight chain (n-16) and cyclic (17:) fatty acids. Based on its morphological, physiological and chemotaxonomic characteristics, this organism was placed in the genusSerratia. Nutritional factors affecting enzyme production were explored. Xylose and ammonium sulfate were the best carbon and nitrogen sources, respectively. Ferrous ions exerted a considerable positive effect on enzyme production. The optimal pH and temperature for phospholipase A1 production were 7.0 and 30°C, respectively. 相似文献
29.
Summary The direct, lipase-catalyzed esterifications of glycerol-3-phosphate in an organic solvent system and in a solvent free system were carried out. In a solvent free system only, LPA synthesis could be achieved within the acceptable reaction time. Open reaction system was preferable to closed reaction system for LPA synthesis. Yield of LPA isolated by silica gel column chromatography was 32.3%. 相似文献
30.
Kinetics of cell death and the production of dissolved organic carbon (DOC) were investigated in Anabaena flos-aquae (Lyngb.) Bréb grown on three different N sources (N2nitrate, and ammonium) in a phosphorus (P)-limited chemostat. The fraction of live cells in the total population increased as growth rate increased with decreasing P limitation. Cell death was less in nitrate and ammonium media than in N2. The specific death rate (γ), when calculated as the slope ofv?1x vs. D?1, where vxand D are live cell fraction (or cell viability) and dilution rate, respectively, was 0. 0082 day?1 in N2and 0.0042 day?1 in nitrate. The slope of the plot in ammonium culture was not significant; however, the value of the live cell fraction was within the range for the NO?3culture. The fraction of live vegetative cells in N2 culture was constant at all growth rates and the increase in the overall live cell fraction with growth rate was due entirely to an increase in live heterocysts. Live heterocysts comprised 3.5% of the total cells at a growth rate of 0.25 day?1 and increased to 6.3% at 0.75 day?1 with the ratio of live heterocysts to live vegetative cells linearly increasing with growth rate. The fraction of live vegetative cells was invariant in nitrate cultures us in N2cultures. The live heterocysts fraction also increased with growth rate in nitrate cultures, along with the live heterocysts : live vegetative cells ratio, but the level was lower than in N2cultures. DOC released from dead cells increased inversely with growth rate in N2from 36.4% of the total DOC at a growth rate of 0.75 day?1 to 54.15% at 0.25 day?1. The contribution of cell death to the total DOC production in nitrate and ammonium media was significantly less than that under N2DOC from dead cells consisted mainly of high-molecular-weight compounds, whereas DOC excreted from live cells was largely of low molecular weight. 相似文献