全文获取类型
收费全文 | 4952篇 |
免费 | 407篇 |
国内免费 | 109篇 |
专业分类
5468篇 |
出版年
2023年 | 37篇 |
2022年 | 83篇 |
2021年 | 174篇 |
2020年 | 128篇 |
2019年 | 163篇 |
2018年 | 199篇 |
2017年 | 138篇 |
2016年 | 188篇 |
2015年 | 258篇 |
2014年 | 291篇 |
2013年 | 389篇 |
2012年 | 399篇 |
2011年 | 336篇 |
2010年 | 223篇 |
2009年 | 189篇 |
2008年 | 243篇 |
2007年 | 208篇 |
2006年 | 215篇 |
2005年 | 186篇 |
2004年 | 172篇 |
2003年 | 161篇 |
2002年 | 157篇 |
2001年 | 132篇 |
2000年 | 98篇 |
1999年 | 97篇 |
1998年 | 51篇 |
1997年 | 35篇 |
1996年 | 36篇 |
1995年 | 28篇 |
1994年 | 15篇 |
1993年 | 21篇 |
1992年 | 30篇 |
1991年 | 26篇 |
1990年 | 23篇 |
1989年 | 27篇 |
1988年 | 25篇 |
1987年 | 24篇 |
1986年 | 18篇 |
1985年 | 25篇 |
1984年 | 20篇 |
1983年 | 12篇 |
1982年 | 12篇 |
1979年 | 16篇 |
1978年 | 15篇 |
1977年 | 14篇 |
1975年 | 11篇 |
1974年 | 10篇 |
1973年 | 11篇 |
1972年 | 12篇 |
1971年 | 12篇 |
排序方式: 共有5468条查询结果,搜索用时 0 毫秒
11.
Odile Gateau Maria Rocha de Morillo Pierre Louisot Renée Morelis 《生物化学与生物物理学报:生物膜》1980,595(1):157-160
A sialyl transferase activity is found in purified mitochondria. It is not due to residual contamination and this enzymatic system is located in the outer mitochondrial membrane. This proves mitochondrial autonomy in regard to glycoconjugate sialylation. 相似文献
12.
Immunoaffinity purification and partial amino acid sequence analysis of catechol-O-methyltransferase from pig liver 总被引:1,自引:0,他引:1
B Bertocci G Garotta M Da Prada H W Lahm G Zürcher G Virgallita V Miggiano 《Biochimica et biophysica acta》1991,1080(2):103-109
Monoclonal antibodies (mAbs) against the soluble form (S-COMT) of catechol-O-methyltransferase (COMT, EC 2.1.1.6) were produced using a purified preparation of the enzyme from pig liver as antigen. The selected monoclonal antibodies recognized the enzyme with different capacities. One of them (Co60-1B/7) showed a significant cross reaction with S-COMT from rat and human liver. A protein band of 23 kDa was recognized by the mAbs on Western blots of the soluble fraction of pig liver. The mAbs were also able to recognize the membrane-bound form of the enzyme, which was found to be mainly localized in the microsomal fraction of pig and rat liver as well as of the human hepatoma cell line Hep G2. The protein bands detected in microsomes had a molecular mass of 26 kDa in pig and rat liver and displayed a slightly higher molecular mass (29 kDa) in the Hep G2 cell line. A single step method for the immunoaffinity purification of pig liver S-COMT was developed by using a Sepharose 4B column to which the mAb Co54-5F/8 was covalently coupled. Acid elution conditions were optimized to obtain the enzyme in active form with a good yield. SDS-PAGE analysis of the purified preparation revealed a single protein band with a molecular mass of 23 kDa with 154-fold enrichment in enzyme activity over the starting material. Since the N-terminus was blocked, purified enzyme preparations were cleaved with trypsin. Two fragments of 22 and 33 amino acids in length could be sequenced by Edman degradation. 相似文献
13.
Influence of dilution rate and induction of cloned gene expression in continuous fermentations of recombinant yeast 总被引:1,自引:0,他引:1
The effects of growth rate on cloned gene product synthesis in recombinant Saccharomyces cerevisiae have been studied in continuous culture. The plasmid employed contains a yeast GAL10-CYC1 hybrid promoter directing expression of the E. coli lacZ gene. beta-Galactosidase production was therefore controlled by the yeast galactose regulatory circuit, and the induction process and its effects were studied at the various dilution rates. At all dilution rates plasmid stability decreased with induction of lacZ gene expression. In some instances, two induced "steady states" were observed, the first 10-15 residence times after induction and the second after 40-50 residence times. The second induced steady state was characterized by greater biomass concentration and lower beta-galactosidase specific activity relative to the first induced "steady-state." beta-Galactosidase specific activity and biomass concentration increased as dilution rate was reduced, and despite lower flow rate and plasmid stability, overall productivity (activity/L/hr) was substantially higher at low dilution rate. Important factors influencing all of the trends were the glucose and galactose (inducer) concentrations in the vessel and inducer metabolism. 相似文献
14.
Influence of the carboxyl terminus of luteinizing hormone-releasing hormone and bradykinin on hydrolysis by brain endo-oligopeptidases 总被引:1,自引:0,他引:1
A C De Camargo M J Da Fonseca H Caldo K De Morais Carvalho 《The Journal of biological chemistry》1982,257(16):9265-9267
A homogeneous preparation of endo-oligopeptidase A from rabbit brain cleaves luteinizing hormone-releasing hormone (less than Glu-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH2) at the Tyr-Gly bond only after the removal of Gly-NH2 from the COOH-terminal position of the molecule. The influence of the carboxyl terminus on hydrolysis by brain endo-oligopeptidases was studied using bradykinin as a model substrate. The substitution of the carboxyl group of bradykinin by the amide reduces by 2.5-fold the rate of Phe-Ser bond hydrolysis by endo-oligopeptidase A but has no effect on the rate of hydrolysis of the Pro-Phe bond by endo-oligopeptidase B. On the other hand, the deletion of Phe-Arg from the COOH-terminal portion of bradykinin makes the peptide resistant to hydrolysis by endo-oligopeptidase A whereas it increases by 5-fold the rate of hydrolysis of the Pro-Gly bond by endo-oligopeptidase B. 相似文献
15.
Adenylate cyclase (ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1) activity in Blastocladiella emersonii is associated with particulate subcellar fractions. Solubilization after treatment with detergent suggests its localization in a membrane fraction of the zoospore homogenate. The enzyme specifically requires Mn2+ for activity and is not stimulated by NaF. The kinetic characteristics of substrate utilization by B. emersonii adenylate cyclase were investigated with various concentrations of ATP and Mn2+, and in the presence of inhibitors. Plots of enzyme activity versus the actual concentration of the MnATP2- complex give sigmoid curves. An excess of Mn2+ activates the enzyme at low concentrations of substrate and leads to a modification of the enzyme kinetics. The nucleotides 5'-AMP and GTP were shown to be competitive inhibitors of the enzyme. In addition, kinetic data, obtained under conditions in which an inhibitor (ATP) is added in constant proportion to the variable substrate (MnATP2-) concentration, produced reciprocal plots that were linear and intersecting to the right of the ordinate, and secondary replots that were hyperbolic. These kinetic patterns support a model in which: MnATP2- is the substrate; free Mn2+ is an activator at low substrate concentrations, but an inhibitor at high substrate concentrations; and free ATP is not an efficient inhibiyor (Ki greater than 1.10(-4) M). 相似文献
16.
Rita Rocha Celso M. Teixeira-Duarte João M.P. Jorge João Henrique Morais-Cabral 《Journal of structural biology》2019,205(3):34-43
RCK (regulating conductance of K+) domains are common regulatory domains that control the activity of eukaryotic and prokaryotic K+ channels and transporters. In bacteria these domains play roles in osmoregulation, regulation of turgor and membrane potential and in pH homeostasis. Whole-genome sequencing unveiled RCK gene redundancy, however the biological role of this redundancy is not well understood. In Bacillus subtilis, there are two closely related RCK domain proteins (KtrA and KtrC) that regulate the activity of the Ktr cation channels. KtrA has been well characterized but little is known about KtrC. We have characterized the structural and biochemical proprieties of KtrC and conclude that KtrC binds ATP and ADP, just like KtrA. However, in solution KtrC exist in a dynamic equilibrium between octamers and non-octameric species that is dependent on the bound ligand, with ATP destabilizing the octameric ring relative to ADP. Accordingly, KtrC-ADP crystal structures reveal closed octameric rings similar to those in KtrA, while KtrC-ATP adopts an open assembly with RCK domains forming a super-helix. In addition, both KtrC-ATP and -ADP octamers are stabilized by the signaling molecule cyclic-di-AMP, which binds to KtrC with high affinity. In contrast, c-di-AMP binds with 100-fold lower affinity to KtrA. Despite these differences we show with an E. coli complementation assay that KtrC and KtrA are interchangeable and able to form functional transporters with both KtrB and KtrD. The distinctive properties of KtrC, in particular ligand-dependent assembly/disassembly, suggest that this protein has a specific physiological role that is distinct from KtrA. 相似文献
17.
Lipase immobilisation on to polymeric membranes 总被引:2,自引:0,他引:2
M. G. Carneiro-da-Cunha J. M. S. Rocha F. A. P. Garcia M. H. Gil 《Biotechnology Techniques》1999,13(6):403-409
Lipase (EC 3.1.1.3) from Candida rugosa was covalently immobilised on to cellulose, cellulose derivatives (cellulose acetate and cellulose phthalate) and cellulose composite membranes using activating agents such as sodium periodate or carbodiimide. Other non-cellulosic polymeric membranes (nylon, polyurethane, chitosan and hydroxyethyl methacrylate-co-methyl methacrylate) were also prepared and used for lipase immobilisation. The results obtained showed that the expressed activities are of the same order of magnitude for similar enzyme loadings when compared with those obtained from literature. 相似文献
18.
19.
A. Braga P. Ferreira J. Oliveira I. Rocha N. Faria 《World journal of microbiology & biotechnology》2018,34(8):122
The polyphenol resveratrol (3,5,4′-trihydroxystilbene) is a well-known plant secondary metabolite, commonly used as a medical ingredient and a nutritional supplement. Due to its health-promoting properties, the demand for resveratrol is expected to continue growing. This stilbene can be found in different plants, including grapes, berries (blackberries, blueberries and raspberries), peanuts and their derived food products, such as wine and juice. The commercially available resveratrol is usually extracted from plants, however this procedure has several drawbacks such as low concentration of the product of interest, seasonal variation, risk of plant diseases and product stability. Alternative production processes are being developed to enable the biotechnological production of resveratrol by genetically engineering several microbial hosts, such as Escherichia coli, Corynebacterium glutamicum, Lactococcus lactis, among others. However, these bacterial species are not able to naturally synthetize resveratrol and therefore genetic modifications have been performed. The application of emerging metabolic engineering offers new possibilities for strain and process optimization. This mini-review will discuss the recent progress on resveratrol biosynthesis in engineered bacteria, with a special focus on the metabolic engineering modifications, as well as the optimization of the production process. These strategies offer new tools to overcome the limitations and challenges for microbial production of resveratrol in industry. 相似文献
20.
Carlos E. Da Costa Joo V. Comasseto Iracema H.-S. Crusius Leandro H. Andrade Andr L.M. Porto 《Journal of Molecular Catalysis .B, Enzymatic》2007,45(3-4):135-139
Aspergillus terreus CCT 3320 and A. terreus URM 3571 catalysed the biotransformations of organic β-hydroxyphenyl selenides through oxidation and methylation reactions. The kinetic resolution of (RS)-1-(phenylseleno)-2-propanol (1) via enantioselective oxidation produced (+)-(S)-1 in high enantiomeric excess (>99%) and in a yield of 50% as determined by product isolation. Oxidation of the R-enantiomer of 1, followed by elimination of the propyl moiety and subsequent methylation of the presumed intermediate, led to the formation of methylphenyl-selenide, which was isolated in a yield of 40%. Whole cells of A. terreus also biocatalysed transformations of diphenyldiselenide, benzeneseleninic acid, (RS)-1-(phenylseleno)-2-pentanol and (RS)-1-(phenylseleno)-3-methyl-2-butanol, but not of (RS)-1-(phenylseleno)-2-phenyl-methanol. This is the first report of the biomethylation of organoselenium compounds by whole cells of A. terreus. 相似文献