全文获取类型
收费全文 | 110486篇 |
免费 | 13242篇 |
国内免费 | 412篇 |
专业分类
124140篇 |
出版年
2018年 | 990篇 |
2017年 | 964篇 |
2016年 | 1258篇 |
2015年 | 1550篇 |
2014年 | 1942篇 |
2013年 | 2462篇 |
2012年 | 2859篇 |
2011年 | 2810篇 |
2010年 | 1884篇 |
2009年 | 1845篇 |
2008年 | 2360篇 |
2007年 | 2343篇 |
2006年 | 2332篇 |
2005年 | 2132篇 |
2004年 | 2071篇 |
2003年 | 2103篇 |
2002年 | 2073篇 |
2001年 | 9640篇 |
2000年 | 9509篇 |
1999年 | 7195篇 |
1998年 | 1614篇 |
1997年 | 1792篇 |
1996年 | 1580篇 |
1995年 | 1407篇 |
1994年 | 1308篇 |
1993年 | 1249篇 |
1992年 | 4792篇 |
1991年 | 4522篇 |
1990年 | 3992篇 |
1989年 | 4012篇 |
1988年 | 3608篇 |
1987年 | 3084篇 |
1986年 | 2776篇 |
1985年 | 2678篇 |
1984年 | 1973篇 |
1983年 | 1729篇 |
1982年 | 1230篇 |
1981年 | 983篇 |
1980年 | 913篇 |
1979年 | 1762篇 |
1978年 | 1361篇 |
1977年 | 1199篇 |
1976年 | 1028篇 |
1975年 | 1149篇 |
1974年 | 1166篇 |
1973年 | 1162篇 |
1972年 | 1033篇 |
1971年 | 955篇 |
1970年 | 822篇 |
1969年 | 743篇 |
排序方式: 共有10000条查询结果,搜索用时 15 毫秒
961.
Purification and properties of a vanadate- and N-ethylmaleimide-sensitive ATPase from chromaffin granule membranes 总被引:2,自引:0,他引:2
A vanadate- and N-ethylmaleimide-sensitive ATPase was purified about 500-fold from chromaffin granule membranes. The purified preparation contained a single major polypeptide with an apparent molecular mass of about 115 kDa, which was copurified with the ATPase activity. Immunological studies revealed that this polypeptide has no relation to subunit I (115 kDa) of the H+-ATPase from chromaffin granules. The ATPase activity of the enzyme is inhibited about 50% by 100 microM N-ethylmaleimide or 5 microM vanadate. The enzyme is not sensitive to dicyclohexylcarbodiimide, ouabain, SCH28080, and omeprazole, which distinguishes it from Na+/K+-ATPase and the gastric K+/H+-ATPase. ATP and 2-deoxy ATP are equally effective substrates for the enzyme. However, the enzyme exhibited only 10% activity with GTP as a substrate. UV illumination of the purified enzyme in the presence of [alpha-32P]ATP exclusively labeled the 115 kDa protein. This labeling was increased by Mg2+ and strongly inhibited by Ca2+ ions. Similarly, the ATPase activity was dependent on Mg2+ and inhibited by the presence of Ca2+ ions. The ATPase activity of the enzyme was largely insensitive to monovalent anions and cations, except for F-, which inhibited the vanadate-sensitive ATPase. Incubation of the enzyme in the presence of [14C]N-ethylmaleimide labeled the 115-kDa polypeptide, and this labeling could be prevented by the addition of ATP during the incubation. A reciprocal experiment showed that preincubation with N-ethylmaleimide inhibited the labeling of the 115-kDa polypeptide by [alpha-32P]ATP by UV illumination. This suggests a close proximity between the ATP-binding site and an essential sulfhydryl group. A possible connection between the isolated ATPase and organelle movement is discussed. 相似文献
962.
963.
Identification of cDNA clones encoding different domains of the basement membrane heparan sulfate proteoglycan 总被引:16,自引:0,他引:16
D M Noonan E A Horigan S R Ledbetter G Vogeli M Sasaki Y Yamada J R Hassell 《The Journal of biological chemistry》1988,263(31):16379-16387
We have used antibodies to the basement membrane proteoglycan to screen lambda gt11 expression vector libraries and have isolated two cDNA clones, termed BPG 5 and BPG 7, which encode different portions of the core protein of the heparan sulfate basement membrane proteoglycan. These clones hybridize to a single mRNA species of approximately 12 kilobases. Amino acid sequences obtained on peptides derived from protease digests of the core protein were found in the deduced sequence, confirming the identity of these clones. BPG 5 spanned 1986 base pairs and has an open reading frame of 662 amino acids. The amino acid sequence deduced from BPG 5 contains two cysteine-rich domains and two internally homologous domains lacking cysteine. The cysteine-rich domains show homology to the cysteine-rich domains of the laminin chains. A globule-rod structure, similar to that of the short arms of the laminin chains, is proposed for this region of the proteoglycan. The other clone, BPG 7, is 2193 base pairs long and has an open reading frame of 731 amino acids. The deduced sequence contains eight internal repeats with 2 cysteine residues in each repeat. These repeats show homology to the neural-cell adhesion molecule N-CAM and the plasma alpha 1B-glycoprotein. Looping structures similar to these proteins and to other proteins of the immunoglobulin gene superfamily are proposed for this region of the proteoglycan. The sequence DSGEY was found four times in this domain and could be heparan sulfate attachment sites. 相似文献
964.
Animal viruses are able to fuse with prokaryotic cells. Fusion between Sendai or influenza virions and Mycoplasma 总被引:2,自引:0,他引:2
V Citovsky S Rottem O Nussbaum Y Laster R Rott A Loyter 《The Journal of biological chemistry》1988,263(1):461-467
Sendai and influenza virions are able to fuse with mycoplasmata. Virus-Mycoplasma fusion was demonstrated by the use of fluorescently labeled intact virions and fluorescence dequenching, as well as by electron microscopy. A high degree of fusion was observed upon incubation of both virions with Mycoplasma gallisepticum or Mycoplasma capricolum. Significantly less virus-cell fusion was observed with Acholeplasma laidlawii, whose membrane contains relatively low amounts of cholesterol. The requirement of cholesterol for allowing virus-Mycoplasma fusion was also demonstrated by showing that a low degree of fusion was obtained with M. capricolum, whose cholesterol content was decreased by modifying its growth medium. Fluorescence dequenching was not observed by incubating unfusogenic virions with mycoplasmata. Sendai virions were rendered nonfusogenic by treatment with trypsin, phenylmethylsulfonyl fluoride, or dithiothreitol, whereas influenza virions were made nonfusogenic by treatment with glutaraldehyde, ammonium hydroxide, high temperatures, or incubation at low pH. Practically no fusion was observed using influenza virions bearing uncleaved hemagglutinin. Trypsinization of influenza virions bearing uncleaved hemagglutinin greatly stimulated their ability to fuse with Mycoplasma cells. Similarly to intact virus particles, also reconstituted virus envelopes, bearing the two viral glycoproteins, fused with M. capricolum. However, membrane vesicles, bearing only the viral binding (HN) or fusion (F) glycoproteins, failed to fuse with mycoplasmata. Fusion between animal enveloped virions and prokaryotic cells was thus demonstrated. 相似文献
965.
Erythrocyte membranes from goat contain a considerable amount, more than 10% of the total amount, of a glycoprotein with Mr = 155,000 (gp155) on sodium dodecyl sulfate-polyacrylamide electrophoresis gel. This report describes the first isolation and characterization of gp155. This gp155 has major trypsin-sensitive sites at each side of the plasma membrane to generate membrane-bound fragments, indicating that the gp155 spans the lipid bilayer several times. This protein consists of a single polypeptide containing about 1,200 amino acid residues corresponding to Mr = 134,000 and some complex type N-linked oligosaccharide chains. A fraction (15-20%) of the gp155 is recovered in nonionic detergent-extracted ghosts along with 25-30% of band 3 and other cytoskeletal proteins and is completely released into solution by extraction with 1 M KCl. Immunoprecipitation with anti-gp155 and anti-ankyrin antibodies of detergent-solubilized membranes separated on a gel permeation chromatography column showed that a part of the gp155 is tightly linked to band 3 with a molar ratio of 1:2 to 1:3. This gp155-band 3 complex in turn is associated to ankyrin through the binding of band 3 to ankyrin. These data indicate that, in native erythrocyte membranes, as well as in detergent solution, gp155 could play a physiological role in controlling cellular integrity and elasticity by forming the gp155-band 3-ankyrin complex. Partial amino acid sequences of the tryptic peptides are also determined. 相似文献
966.
Purification and identification of [hydroxyprolyl3]bradykinin in ascitic fluid from a patient with gastric cancer 总被引:3,自引:0,他引:3
Kinins in the ascitic fluid from a patient with gastric cancer were purified by gel filtration and reversed-phase high-performance liquid chromatography (HPLC). Two fractions (fractions I and II) showed kinin activity. Fraction I did not correspond to either bradykinin or other known kinins, whereas fraction II corresponded to bradykinin. Fraction I contained 8 amino acid residues from bradykinin minus 1 proline plus 1 additional hydroxyproline. Sequence analysis of fraction I showed that the proline at the third amino acid residue of bradykinin was replaced by hydroxyproline. The retention time of fraction I on reversed-phase HPLC was exactly the same as that of synthetic [hydroxyprolyl3]bradykinin (Arg-Pro-Hyp-Gly-Phe-Ser-Pro-Phe-Arg) and was distinguishable from des-Pro3-bradykinin. Thus, these results demonstrate for the first time the presence of [hydroxyprolyl3]bradykinin in vivo. This is also the first report of the presence of bradykinin in human tumor ascites. 相似文献
967.
Specificity and mechanism of influence of amino acid residues on hepatic clearance of oligopeptides 总被引:2,自引:0,他引:2
We have investigated the influence of amino acid residues on hepatic clearance of oligopeptides by determining the rate of disappearance (nmol.(min.g liver)-1) of selective oligopeptides from the medium during isolated rat liver perfusion. (a) N terminus: the rate of disappearance of Ala-Leu was greater (p less than 0.01) than those of Gly-Leu, Phe-Leu, and Arg-Leu (208 +/- 13, 135 +/- 13, 116 +/- 12, and 127 +/- 12, respectively). (b) C terminus: the rate of disappearance of Leu-Ala (244 +/- 18) was significantly greater (p less than 0.01) than that of Leu-Gly (145 +/- 16). (c) Number of residues: with each increase in the number of alanine residues (2-4) there was a significant increase in the rate of peptide disappearance, and conversely, with each increase in the number of glycine residues (2-6) there was a significant decrease in the rate of peptide disappearance. Further studies showed no peptide transport by isolated liver plasma membrane vesicles and no significant correlation between the rates of peptide disappearance and hydrolase activities of the perfusion medium but highly significant correlation with hydrolase activity of plasma membrane. We conclude that certain amino acid residues, such as alanine, enhance hepatic clearance of oligopeptides by increasing their affinity as substrates for plasma membrane peptide hydrolases. 相似文献
968.
Structure, biosynthesis, and glycosylation of human small intestinal maltase-glucoamylase 总被引:1,自引:0,他引:1
Maltase-glucoamylase (MGA) was immunoprecipitated from detergent extracts of brush border membranes of the human small intestinal mucosa. Electrophoretic analysis of the precipitates under denaturing conditions revealed a single polypeptide of Mr = 335,000 in the presence or absence of reducing agents. Cross-linking of brush border membranes with the homobifunctional reagent dithiobis(succinimidylpropionate) did not result in considerable changes in the electrophoretic pattern of MGA. In contrast, aminopeptidase N, used in these studies as a control glycoprotein of the brush border membrane revealed dimeric structures of its single subunit in the presence of dithiobis(succinimidylpropionate). These data suggest that MGA is expressed in the human small intestinal brush border as a monomeric polypeptide. The biosynthesis of MGA was studied by pulse-labeling of human intestinal biopsy specimens or mucosal explants in organ culture. Continuous labeling with [35S]methionine for 30 min revealed a single polypeptide high mannose precursor of Mr = 285,000 (MGAh) which matures after 4 h of labeling to the Mr = 335,000 as judged by the susceptibility of these two forms to endo-beta-N-acetylglucosaminidase H. Owing to the absence of pancreatic secretions in the culture medium and the isolation of an identical species from nonlabeled mucosa, this result indicates that the Mr = 335,000 does not undergo an in situ extracellular cleavage by intraluminal proteases. Further, biosynthetically labeled, intracellularly cleaved polypeptides corresponding to the high mannose precursor or mature forms of MGA were not detected. The mature form of MGA (MGAm) bears in addition to N-linked glycans also O-glycosidically linked oligosaccharides. In fact, endo-beta-N-acetylglucosaminidase F/glycopeptidase F treatment of MGAm followed by chemical deglycosylation with trifluoromethanesulfonic acid revealed approximately 35,000 daltons of O-linked sugars. Furthermore, MGAm as well as its N-linked sugars-depleted form bound to Helix pomatia lectin which has specificity toward Gal-GalNAc structures. In addition, the data were suggestive of a post-translational O-glycosylation of the molecule since (i) the high mannose precursor of MGA did not bind to H. pomatia lectin and (ii) its endo-beta-N-acetylglucosaminidase H or endo-beta-N-acetylglucosaminidase F/glycopeptidase F form displayed an apparent molecular weight similar to that obtained upon endo-beta-N-acetylglucosaminidase F/glycopeptidase F/trifluoromethanesulfonic acid deglycosylation. Finally, pulse-chase experiments revealed a relatively slow rate of post-translational processing of MGA in comparison to aminopeptidase N.(ABSTRACT TRUNCATED AT 400 WORDS) 相似文献
969.
The activities of cofactor-independent and CoA-dependent transacylation were examined for various rabbit tissues. Liver microsomes were found to exhibit relatively high CoA-dependent transacylation activity, while the cofactor-independent transacylation activity was low. The apparent Km values for CoA were 1.4 microM (acceptor, 1-acyl-sn-glycero-3-phosphocholine (1-acyl-GPC] and 3.8 microM (acceptor, 1-acyl-sn-glycero-3-phosphoethanolamine (1-acyl-GPE], respectively. The apparent Vmax values were 2.6 nmol/min/mg (1-acyl-GPC) and 1.2 nmol/min/mg (1-acyl-GPE), respectively. The CoA-dependent transacylation reaction shows a distinct fatty acid specificity. [14C]18:2 and [14C]20:4 at the 2-positions and [14C]18:0 at the 1-positions of donor phospholipids were transferred to lysophospholipids in the presence of CoA. We observed the formation of considerable amounts of acyl-CoA from these fatty acids during the reaction, without the participation of ATP. The transfer of other fatty acids between phospholipids was shown to be almost nil. The very low transfer of 18:1 was in marked contrast to the effective utilization of 18:1-CoA by acyl-CoA:1-acyl-GPC acyltransferase. The effects of several compounds and heat treatment on these two acylation reactions were also examined. The CoA-dependent transacylation reaction may be important for the selective acylation of certain lysophospholipids, such as 1-acyl-GPE, in living cells with the cooperation of acyl-CoA:lysophospholipid acyltransferase, which generates CoA for the former reaction. 相似文献
970.
Structure of an extracellular cross-reactive polysaccharide from Pseudomonas aeruginosa immunotype 4 总被引:6,自引:0,他引:6
N A Kocharova Y A Knirel A S Shashkov N K Kochetkov G B Pier 《The Journal of biological chemistry》1988,263(23):11291-11295
A neutral small molecular mass (approximately 6.5 kDa) polysaccharide comprising a pentasaccharide repeat unit was isolated from culture supernatants of Pseudomonas aeruginosa immunotype 4. The polysaccharide had a pentasaccharide repeating unit as follows (formula; see text) where Rha is rhamnose. The structure was determined using acid hydrolysis, solvolysis with anhydrous hydrogen fluoride, methylation analysis, and 1H and 13C nuclear magnetic resonance spectroscopy including nuclear Overhauser enhancement experiments. The polysaccharide bound antibody raised to the lipopolysaccharide of the seven P. aeruginosa Fisher-Devlin immunotype strains. Inhibition assays demonstrated the presence of a serologically similar polysaccharide in supernatants of these strains. Affinity-purified antibody to the polysaccharide bound to lipopolysaccharide and whole cells of the immunotype strains of P. aeruginosa in a Western immunoblot and colony blot assay, respectively. This polysaccharide seems to contain an antigenic determinant present in the core of the P. aeruginosa lipopolysaccharide or may represent another minor polysaccharide substituent on the lipopolysaccharide in addition to the O side chain. 相似文献