全文获取类型
收费全文 | 1002篇 |
免费 | 131篇 |
国内免费 | 75篇 |
专业分类
1208篇 |
出版年
2024年 | 3篇 |
2023年 | 10篇 |
2022年 | 25篇 |
2021年 | 40篇 |
2020年 | 33篇 |
2019年 | 39篇 |
2018年 | 38篇 |
2017年 | 40篇 |
2016年 | 48篇 |
2015年 | 62篇 |
2014年 | 72篇 |
2013年 | 61篇 |
2012年 | 104篇 |
2011年 | 80篇 |
2010年 | 46篇 |
2009年 | 46篇 |
2008年 | 48篇 |
2007年 | 27篇 |
2006年 | 45篇 |
2005年 | 33篇 |
2004年 | 39篇 |
2003年 | 49篇 |
2002年 | 34篇 |
2001年 | 36篇 |
2000年 | 25篇 |
1999年 | 25篇 |
1998年 | 12篇 |
1997年 | 10篇 |
1996年 | 11篇 |
1995年 | 9篇 |
1994年 | 4篇 |
1993年 | 8篇 |
1992年 | 14篇 |
1991年 | 7篇 |
1990年 | 5篇 |
1989年 | 1篇 |
1988年 | 4篇 |
1987年 | 6篇 |
1986年 | 1篇 |
1985年 | 3篇 |
1984年 | 2篇 |
1983年 | 1篇 |
1982年 | 2篇 |
排序方式: 共有1208条查询结果,搜索用时 15 毫秒
981.
982.
Mikhail O. Maksimov Joseph D. Koos Chuhan Zong Bozhena Lisko A. James Link 《The Journal of biological chemistry》2015,290(52):30806-30812
Lasso peptide isopeptidase is an enzyme that specifically hydrolyzes the isopeptide bond of lasso peptides, rendering these peptides linear. To carry out a detailed structure-activity analysis of the lasso peptide isopeptidase AtxE2 from Asticcacaulis excentricus, we solved NMR structures of its substrates astexin-2 and astexin-3. Using in vitro enzyme assays, we show that the C-terminal tail portion of these peptides is dispensable with regards to isopeptidase activity. A collection of astexin-2 and astexin-3 variants with alanine substitutions at each position within the ring and the loop was constructed, and we showed that all of these peptides except for one were cleaved by the isopeptidase. Thus, much like the lasso peptide biosynthetic enzymes, lasso peptide isopeptidase has broad substrate specificity. Quantitative analysis of the cleavage reactions indicated that alanine substitutions in loop positions of these peptides led to reduced cleavage, suggesting that the loop is serving as a recognition element for the isopeptidase. 相似文献
983.
Hai‐Yang Wang Ye Yao Qing‐Feng Sun Zong‐Hong Liu Si‐Qi Liu Jun‐Li Zhuang Yun‐Peng Wang Hong‐Yu Liu 《Journal of cellular and molecular medicine》2017,21(8):1482-1491
This study aimed to investigate the role of miR‐138 in human coronary artery endothelial cell (HCAEC) injury and inflammatory response and the involvement of the PI3K/Akt/eNOS signalling pathway. Oxidized low‐density lipoprotein (OX‐LDL)‐induced HCAEC injury models were established and assigned to blank, miR‐138 mimic, miR‐138 inhibitor, LY294002 (an inhibitor of the PI3K/Akt/eNOS pathway), miR‐138 inhibitor + LY294002 and negative control (NC) groups. qRT‐PCR and Western blotting were performed to detect the miR‐138, PI3K, Akt and eNOS levels and the protein expressions of PI3K, Akt, eNOS, p‐Akt, p‐eNOS, Bcl‐2, Bax and caspase‐3. ELISAs were employed to measure the expressions of TNF‐α, IL‐4, IL‐6, IL‐8, IL‐10 and nitric oxide (NO) and the activities of lactate dehydrogenase (LDH) and eNOS. MTT and flow cytometry were performed to assess the proliferation and apoptosis of HCAECs. Compared to the blank group, PI3K, Akt and eNOS were down‐regulated in the miR‐138 mimic and LY294002 groups but were up‐regulated in the miR‐138 inhibitor group. The miR‐138 mimic and LY294002 groups showed decreased concentrations of TNF‐α, IL‐6, IL‐8 and NO and reduced activities of LDH and eNOS, while opposite trends were observed in the miR‐138 inhibitor group. The concentrations of IL‐4 and IL‐10 increased in the miR‐138 mimic and LY294002 groups but decreased in the miR‐138 inhibitor group. The miR‐138 mimic and LY294002 groups had significantly decreased cell proliferation and increased cell apoptosis compared to the blank group. These findings indicate that up‐regulation of miR‐138 alleviates HCAEC injury and inflammatory response by inhibiting the PI3K/Akt/eNOS signalling pathway. 相似文献
984.
Producing defucosylated antibodies with enhanced in vitro antibody‐dependent cellular cytotoxicity via FUT8 knockout CHO‐S cells 下载免费PDF全文
Huifang Zong Lei Han Kai Ding Jiaxian Wang Tao Sun Xinyu Zhang Cedric Cagliero Hua Jiang Yueqing Xie Jianrong Xu Baohong Zhang Jianwei Zhu 《Engineering in Life Science》2017,17(7):801-808
To engineer a host cell line that produces defucosylated mAbs with superior antibody‐dependent cellular cytotoxicity, we disrupted α‐1, 6 fucosyltransferase (FUT8 ) gene in CHO‐S (CHO is Chinese hamster ovary) cells by clustered regularly interspaced short palindromic repeats‐CRISPR associated nuclease 9. The gene knockout cell line was evaluated for growth, stability, and product quality. The growth profile of FUT8 gene knockout CHO‐S (FUT8 ?/?) cells was comparable with wild type CHO‐S cells. FUT8 catalyzes the transfer of a fucose residue from GDP‐fucose to N‐glycans residue. Defucosylated IgG1 antibodies produced by FUT8 ?/? cells showed increased binding affinities to human FcγRIIIa and higher activities in mediating antibody‐dependent cellular cytotoxicity, comparing with conventional fucosylated IgG1. Our results demonstrated the potential of using the clustered regularly interspaced short palindromic repeats‐CRISPR associated nuclease 9 technology in cell line engineering for biopharmaceutical industrial applications. 相似文献
985.
986.
The coupling between G protein-coupled receptors (GPCRs) and guanine nucleotide-binding proteins (G proteins) regulates various signal transductions from extracellular space into the cell. However, the coupling mechanism between GPCRs and G proteins is still unknown, and experimental determination of their coupling specificity and function is both expensive and time consuming. Therefore, it is significant to develop a theoretical method to predict the coupling specificity between GPCRs and G proteins as well as their function using their primary sequences. In this study, a novel four-layer predictor (GPCRsG_CWTIT) based on support vector machine (SVM), continuous wavelet transform (CWT) and information theory (IT) is developed to classify G proteins and predict the coupling specificity between GPCRs and G proteins. SVM is used for construction of models. CWT and IT are used to characterize the primary structure of protein. Performance of GPCRsG_CWTIT is evaluated with cross-validation test on various working dataset. The overall accuracy of the G proteins at the levels of class and family is 98.23 and 85.42%, respectively. The accuracy of the coupling specificity prediction varies from 74.60 to 94.30%. These results indicate that the proposed predictor is an effective and feasible tool to predict the coupling specificity between GPCRs and G proteins as well as their functions using only the protein full sequence. The establishment of such an accurate prediction method will facilitate drug discovery by improving the ability to identify and predict protein-protein interactions. GPCRsG_CWTIT and dataset can be acquired freely on request from the authors. 相似文献
987.
Interest in essential genes has arisen recently given their importance in antimicrobial drug development. Although knockouts of essential genes are commonly known to cause lethal phenotypes, there is insufficient understanding on the intermediate changes followed by genetic perturbation and to what extent essential genes correlate to other genes. Here, we characterized the gene knockout effects by using a list of affected genes, termed as 'damage lists'. These damage lists were identified through a refined cascading failure approach that was based on a previous topological flux balance analysis. Using an Escherichia coli metabolic network, we incorporated essentiality information into damage lists and revealed that the knockout of an essential gene mainly affects a large range of other essential genes whereas knockout of a non-essential gene only interrupts other non-essential genes. Also, genes sharing common damage lists tend to have the same essentiality. We extracted 72 core functional modules from the common damage lists of essential genes and demonstrated their ability to halt essential metabolites production. Overall, our network analysis revealed that essential and non-essential genes propagated their deletion effects via distinct routes, conferring mechanistic explanation to the observed lethality phenotypes of essential genes. 相似文献
988.
Nuria Martinez‐Lopez Diana Athonvarangkul Srabani Sahu Luisa Coletto Haihong Zong Claire C Bastie Jeffrey E Pessin Gary J Schwartz Rajat Singh 《EMBO reports》2013,14(9):795-803
Macroautophagy (MA) regulates cellular quality control and energy balance. For example, loss of MA in aP2‐positive adipocytes converts white adipose tissue (WAT) into brown adipose tissue (BAT)‐like, enhancing BAT function and thereby insulin sensitivity. However, whether MA regulates early BAT development is unknown. We report that deleting Atg7 in myogenic Myf5+ progenitors inhibits MA in Myf5‐cell‐derived BAT and muscle. Knock out (KO) mice have defective BAT differentiation and function. Surprisingly, their body temperature is higher due to WAT lipolysis‐driven increases in fatty acid oxidation in ‘Beige’ cells in inguinal WAT, BAT and muscle. KO mice also present impaired muscle differentiation, reduced muscle mass and glucose intolerance. Our studies show that ATG7 in Myf5+ progenitors is required to maintain energy and glucose homeostasis through effects on BAT and muscle development. Decreased MA in myogenic progenitors with age and/or overnutrition might contribute to the metabolic defects and sarcopenia observed in these conditions. 相似文献
989.
热量限制延缓人二倍体成纤维细胞衰老的体外模型 总被引:3,自引:0,他引:3
为建立人二倍体成纤维细胞IMR 90的热量限制体外模型 ,分别采用低浓度、正常浓度和高浓度葡萄糖培养条件 ,常规传代培养IMR 90细胞 ,利用综合细胞衰老指标对模型进行评价 .低、正常和高浓度葡萄糖培养条件组IMR 90细胞平均寿限分别为 5 8 3、5 5 0和 4 7 2PDL(群体倍增水平 ) .低浓度葡萄糖培养IMR 90细胞早期增长速度有所减慢 ,但仍保持对生长因子诱导的细胞增殖能力 ,并使晚期IMR 90处于细胞周期S期的比例以及其DNA修复能力显著高于其他条件培养的晚期细胞 .低浓度葡萄糖培养IMR 90晚期细胞的半乳糖苷酶染色阳性率亦明显低于其他条件培养的晚期细胞 .实验结果表明 ,低浓度葡萄糖培养可以延缓IMR 90复制衰老 ,建立了热量限制延缓衰老体外模型 ,为进一步探讨热量限制延缓衰老作用机制的研究打下基础 相似文献
990.
In this paper, highly regioselective enzymatic acylations of 1-β-D-arabinofuranosylcytosine (ara-C) with vinyl stearate (VS) in binary organic solvents were explored for the preparation
of 5′-O-stearate of ara-C with potential antitumor activity. Twelve kinds of hydrolases were tested for the regioselective acylation
reaction and the immobilized Candida antarctica lipase B (Novozym 435) showed the highest regioselectivity (>99.9%) towards the 5′-OH of ara-C. A comparative study showed
that the lipase had much higher catalytic activity in the binary mixture of hexane and pyridine than in other tested co-solvent
systems. To better understand lipase-mediated acylation conducted in the best binary organic solvent system, the effects of
hydrophobic solvent content, molar ratio of VS to ara-C, initial water activity, and reaction temperature on the acylation
reaction were studied. It was found that the most suitable hexane content, VS–ara-C molar ratio, initial water activity, and
reaction temperature were shown to be 25% (v/v), 20:1, 0.07, and 50°C, respectively. Under these reaction conditions, the initial reaction rate, the maximum substrate conversion,
and regioselectivity were as high as 86.0 mmol·L−1h−1, 96.6%, and >99.9%, respectively. The product of Novozym 435-catalyzed acylation was characterized by Carbon-13(13C) NMR and confirmed to be 5′-O-stearate of ara-C. 相似文献