首页 | 本学科首页   官方微博 | 高级检索  
文章检索
  按 检索   检索词:      
出版年份:   被引次数:   他引次数: 提示:输入*表示无穷大
  收费全文   463篇
  免费   35篇
  国内免费   1篇
  2022年   3篇
  2021年   7篇
  2020年   3篇
  2019年   6篇
  2018年   7篇
  2017年   9篇
  2016年   7篇
  2015年   26篇
  2014年   15篇
  2013年   26篇
  2012年   32篇
  2011年   39篇
  2010年   15篇
  2009年   24篇
  2008年   12篇
  2007年   26篇
  2006年   31篇
  2005年   28篇
  2004年   25篇
  2003年   24篇
  2002年   16篇
  2001年   10篇
  2000年   14篇
  1999年   11篇
  1998年   11篇
  1997年   3篇
  1996年   4篇
  1995年   3篇
  1994年   10篇
  1992年   6篇
  1991年   5篇
  1990年   3篇
  1989年   5篇
  1988年   3篇
  1987年   3篇
  1986年   4篇
  1985年   4篇
  1984年   5篇
  1983年   2篇
  1982年   1篇
  1981年   1篇
  1979年   2篇
  1978年   1篇
  1977年   2篇
  1973年   1篇
  1972年   3篇
  1971年   1篇
排序方式: 共有499条查询结果,搜索用时 15 毫秒
51.
We have previously engineered a recombinant Pichia pastoris GS115 transformant, MSPGA-7, harboring seven copies of glucoamylase (GA) fused with modified signal peptide. High yield secretion of GA was achieved as an extra copy of SEC4 was integrated to the transformant. To elucidate the physiological role of SEC4, a dominant-negative mutant of SEC4, SEC4(S28N), was overexpressed under the control of alchohol oxidase 1 (AOX1) promoter in P. pastoris strain MSPGA-7 as well as a set of host cells harboring multi-copy of wild type SEC4. We found that SEC4(S28N) mutation in the key guanine nucleotide binding domain reduced guanine nucleotide binding affinity, hence it blocked the transport of vesicles required for targeting and fusion to the plasma membrane. The inhibitory levels of cell growth and GA secretion were correlated with the dosage of SEC4(S28N) gene. In addition, overexpression of SEC4 driven by AOX1 promoter in MSPGA-7 improved the secretory production of GA, but demonstrated the delay of cell growth by increased gene dosage of SEC4. Interestingly, a limited level of Sec4p did not disturb the cell growth. It was because expression of only one copy of SEC4 resulted in delay of cell growth at an early stage while still maintaining high level Sec4p at long-term incubation. Accordingly, as glyceraldehyde-3-phosphate dehydrogenase promoter was used to substitute AOX1 promoter to drive the SEC4 expression, enhanced GA secretion but not inhibition of cell growth was achieved. Taken together, our results demonstrate that SEC4 is essential for P. pastoris in regulating cell growth and heterologous protein secretion in a dosage-dependent manner.  相似文献   
52.
Human apolipoprotein E (apoE) is a 299-amino-acid protein with a molecular weight of 34 kDa. The difference between the apoE3 and apoE4 isoforms is a single residue substitution involving a Cys-Arg replacement at residue 112. ApoE4 is positively associated with atherosclerosis and late-onset and sporadic Alzheimer's disease (AD). ApoE4 and its C-terminal truncated fragments have been found in the senile plaques and neurofibrillary tangles in the brain of AD patients. However, detail structural information regarding isoform and domain interaction remains poorly understood. We prepared full-length, N-, and C-terminal truncated apoE3 and apoE4 proteins and studied their structural variation. Sedimentation velocity and continuous size distribution analysis using analytical ultracentrifugation revealed apoE3(72-299) as consisting of a major species with a sedimentation coefficient of 5.9. ApoE4(72-299) showed a wider and more complicated species distribution. Both apoE3 and E4 N-terminal domain (1-191) existed with monomers as the major component together with some tetramer. The oligomerization and aggregation of apoE protein increased when the C-terminal domain (192-271) was incorporated. The structural influence of the C-terminal domain on apoE is to assist self-association with no significant isoform preference. Circular dichroism and fluorescence studies demonstrated that apoE4(72-299) possessed a more alpha-helical structure with more hydrophobic residue exposure. The structural variation of the N-terminal truncated apoE3 and apoE4 protein provides useful information that helps to explain the greater aggregation of the apoE4 isoform and thus has implication for the involvement of apoE4 in AD.  相似文献   
53.
The anticancer activity of anti-bacterial cecropins makes them potentially useful as peptide anti-cancer drugs. We used the cell-attached patch to study the effect of cecropin B (CB; having one hydrophobic and one amphipathic alpha-helix) and its derivative, cecropin B3 (CB3; having two hydrophobic alpha-helices) on the membrane of Ags cancer cells. Application of 10-60 microM CB onto the membrane of the cancer cell produces short outward currents. Comparative study with CB3, which induces no outward currents, shows that the amphipathic group of CB is necessary for the pore formation. The results provide a rationale to study the cell-killing activity of antimicrobial peptides at the single cancer cell level.  相似文献   
54.
Huang YL  Leu SF  Liu BC  Sheu CC  Huang BM 《Life sciences》2004,75(9):1051-1062
Cordyceps sinensis (CS), an Ascomycetes fungus parasitic to Lepidoptera larvae, has been traditionally used as nutritious food for the enhancement on sexual performance and the restitution of impairment in sexual function in Chinese society. We have previously demonstrated the stimulatory effect of CS and its fractions on steroidogenesis both on primary mouse Leydig cells and MA-10 mouse Leydig tumor cells. In the present studies, we determined the in vivo effects of CS and its fractions on steroidogenesis in mouse. Different concentrations of CS and CS fractions (0.02 and 0.2 mg/g body weight) were fed to immature or mature mice from 1 to 7 days. The plasma levels of testosterone were evaluated by radioimmunoassay. The weights of reproductive organs were also determined. Results illustrated that CS significantly induced plasma testosterone levels both in immature and mature mice in 3 and/or 7 days treatment (p < 0.05). F2 and F3 at 0.02 and/or 0.2 mg/g body weight for different feeding duration could also significantly stimulated plasma testosterone levels both in immature and mature mice (p < 0.05). In general, CS, F2 and F3 didn't have considerable effect on the weights of reproductive organs. Taken together, these studies illustrate that CS and its fractions significantly stimulated in vivo mouse testosterone production.  相似文献   
55.
Summary Chronic infections with hepatitis B and C viruses (HBV and HCV) are etiologically linked to hepatitis, liver cirrhosis, and hepatocellular carcinoma (HCC). Both viruses may induce activation of nuclear factor-kappa B (NF-κB) in hepatocytes that plays a crucial role in the regulation of cell growth and apoptosis. Functional proteomics analysis of proteins associated with NF-κB signaling complexes in both viruses-related HCC tumor and non-tumor tissues may disclose possible common mechanisms in hepatocarcinogenesis. By functional proteomics, we analyzed proteins associated with NF-κB-signaling complexes in four-paired human HCC tumor and non-tumor tissues from HBV- and HCV-infected patients, respectively, and in one-paired tissue with dual viral infection. The quantity of NF-κB-associated proteins was semi-quantitatively measured by protein spot intensity on the gels of two-dimensional polyacrylamide gel electrophoresis. The results showed that overexpression of NF-κB-associated Wnt-1 protein in tumor part was detected in the␣majority of HBV- and HCV-infected HCC samples. These data suggest that enhanced expression of NF-κB-associated Wnt-1 protein might be a mechanism of hepatocarcinogenesis common to HBV- and HCV-infected patients. NF-κB signaling pathway and Wnt-1 protein could be potential targets for designing highly effective therapeutic agents in treating HCC and for chemoprevention of hepatocarcinogenesis.  相似文献   
56.
In this study, we investigated the signaling pathway involved in IL-6 production caused by peptidoglycan (PGN), a cell wall component of the Gram-positive bacterium, Staphylococcus aureus, in RAW 264.7 macrophages. PGN caused concentration- and time-dependent increases in IL-6, PGE(2), and cAMP production. PGN-mediated IL-6 production was inhibited by a nonselective cyclooxygenase (COX) inhibitor (indomethacin), a selective COX-2 inhibitor (NS398), a PGE(2) (EP2) antagonist (AH6809), a PGE(4) (EP4) antagonist (AH23848), and a protein kinase A (PKA) inhibitor (KT5720), but not by a nonselective NO synthase inhibitor (N(G)-nitro-l-arginine methyl ester). Furthermore, PGE(2), an EP2 agonist (butaprost), an EP2/PGE(3) (EP3)/EP4 agonist (misoprostol), and misoprostol in the presence of AH6809 all induced IL-6 production, whereas an EP1/EP3 agonist (sulprostone) did not. PGN caused time-dependent activations of IkappaB kinase alphabeta (IKKdbeta) and p65 phosphorylation at Ser(276), and these effects were inhibited by NS398 and KT5720. Both PGE(2) and 8-bromo-cAMP also caused IKKdbeta kinase alphabeta phosphorylation. PGN resulted in two waves of the formation of NF-kappaB-specific DNA-protein complexes. The first wave of NF-kappaB activation occurred at 10-60 min of treatment, whereas the later wave occurred at 2-12 h of treatment. The PGN-induced increase in kappaB luciferase activity was inhibited by NS398, AH6809, AH23848, KT5720, a protein kinase C inhibitor (Ro31-8220), and a p38 MAPK inhibitor (SB203580). These results suggest that PGN-induced IL-6 production involves COX-2-generated PGE(2), activation of the EP2 and EP4 receptors, cAMP formation, and the activation of PKA, protein kinase C, p38 MAPK, IKKdbeta, kinase alphabeta, p65 phosphorylation, and NF-kappaB. However, PGN-induced NO release is not involved in the signaling pathway of PGN-induced IL-6 production.  相似文献   
57.
58.
Accumulated evidence reveals that increased cyclooxygenase-2 (COX-2) is involved in the development of colorectal cancer. Our purpose was to quantitate COX-2 expression in colorectal cancers using tissue microarray analysis and look for an association with clinicopathological stage. Immunohistochemical analysis of COX-2 was performed in tissue microarray slides containing 90 specimens including 32 well-differentiated, 35 moderately differentiated, and 23 poorly differentiated colorectal adenocarcinomas. All colorectal adenocarcinomas showed significant immunohistochemical expression of COX-2 when compared to normal colon epithelia. However, there was no significant difference in immunostaining scores between poorly, moderately, and well-differentiated tumors (195 +/- 28, 214 +/- 26 and 200 +/- 24, respectively). The COX-2 immunostaining score correlated significantly with T stage (P < 0.05) but not with N or M stage. The positive expression rates of CK20 were 97% for well-differentiated, 94% for moderately differentiated, and 65% for poorly differentiated colorectal adenocarcinomas, suggesting that CK20 may not be an effective discriminator between poorly differentiated colorectal adenocarcinoma and metastatic adenocarcinoma.  相似文献   
59.
60.
A bacterial strain designated antisso-27T, previously isolated from saltpan in Taiwan while screening for bacteria for algicidal activity, was characterized using the polyphasic taxonomic approach. Strain antisso-27T was Gram-negative, aerobic, brownish yellow colored, rod-shaped, non-flagellated and non-gliding. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain antisso-27T belonged to the genus Aquimarina within the family Flavobacteriaceae with relatively low sequence similarities of 94.0–96.6% to other valid Aquimarina spp. It contained iso-C17:0 3-OH, iso-C15:0, iso-C16:0, iso-C15:1 and iso-C15:0 3-OH as the main fatty acids and contained a menaquinone with six isoprene units (MK-6) as the major isoprenoid quinone. Major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol, an uncharacterized aminolipid and five uncharacterized phospholipids. Strain antisso-27T employed direct mode of algicidal lysis to Chlorella vulgaris strain 211-31; nevertheless, it released an algicidal substance against M. aeruginosa strain MTY01. This is the first study that the Aquimarina species possesses both direct and indirect algicidal activities. On the basis of the phylogenetic and phenotypic data, strain antisso-27T should be classified as representing a novel species, for which the name A. salinaria sp. nov. is proposed. The type strain is A. salinaria antisso-27T (= BCRC 80080T = LMG 25375T).  相似文献   
设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号