全文获取类型
收费全文 | 2105篇 |
免费 | 170篇 |
国内免费 | 1篇 |
出版年
2024年 | 4篇 |
2023年 | 27篇 |
2022年 | 63篇 |
2021年 | 135篇 |
2020年 | 70篇 |
2019年 | 81篇 |
2018年 | 81篇 |
2017年 | 63篇 |
2016年 | 107篇 |
2015年 | 138篇 |
2014年 | 181篇 |
2013年 | 157篇 |
2012年 | 182篇 |
2011年 | 193篇 |
2010年 | 103篇 |
2009年 | 79篇 |
2008年 | 112篇 |
2007年 | 92篇 |
2006年 | 80篇 |
2005年 | 86篇 |
2004年 | 79篇 |
2003年 | 64篇 |
2002年 | 43篇 |
2001年 | 3篇 |
2000年 | 7篇 |
1999年 | 5篇 |
1998年 | 6篇 |
1997年 | 9篇 |
1996年 | 8篇 |
1995年 | 3篇 |
1994年 | 4篇 |
1993年 | 1篇 |
1992年 | 5篇 |
1991年 | 1篇 |
1990年 | 2篇 |
1982年 | 1篇 |
1978年 | 1篇 |
排序方式: 共有2276条查询结果,搜索用时 672 毫秒
991.
Anirudh K. Singh Benjamin Pluvinage Melanie A. Higgins Ankur B. Dalia Shireen A. Woodiga Matthew Flynn Audrey R. Lloyd Jeffrey N. Weiser Keith A. Stubbs Alisdair B. Boraston Samantha J. King 《PLoS pathogens》2014,10(9)
Bacterial cell-surface proteins play integral roles in host-pathogen interactions. These proteins are often architecturally and functionally sophisticated and yet few studies of such proteins involved in host-pathogen interactions have defined the domains or modules required for specific functions. Streptococcus pneumoniae (pneumococcus), an opportunistic pathogen that is a leading cause of community acquired pneumonia, otitis media and bacteremia, is decorated with many complex surface proteins. These include β-galactosidase BgaA, which is specific for terminal galactose residues β-1–4 linked to glucose or N-acetylglucosamine and known to play a role in pneumococcal growth, resistance to opsonophagocytic killing, and adherence. This study defines the domains and modules of BgaA that are required for these distinct contributions to pneumococcal pathogenesis. Inhibitors of β-galactosidase activity reduced pneumococcal growth and increased opsonophagocytic killing in a BgaA dependent manner, indicating these functions require BgaA enzymatic activity. In contrast, inhibitors increased pneumococcal adherence suggesting that BgaA bound a substrate of the enzyme through a distinct module or domain. Extensive biochemical, structural and cell based studies revealed two newly identified non-enzymatic carbohydrate-binding modules (CBMs) mediate adherence to the host cell surface displayed lactose or N-acetyllactosamine. This finding is important to pneumococcal biology as it is the first adhesin-carbohydrate receptor pair identified, supporting the widely held belief that initial pneumococcal attachment is to a glycoconjugate. Perhaps more importantly, this is the first demonstration that a CBM within a carbohydrate-active enzyme can mediate adherence to host cells and thus this study identifies a new class of carbohydrate-binding adhesins and extends the paradigm of CBM function. As other bacterial species express surface-associated carbohydrate-active enzymes containing CBMs these findings have broad implications for bacterial adherence. Together, these data illustrate that comprehending the architectural sophistication of surface-attached proteins can increase our understanding of the different mechanisms by which these proteins can contribute to bacterial pathogenesis. 相似文献
992.
Marcy R. Auerbach Donghong Yan Rajesh Vij Jo-Anne Hongo Gerald Nakamura Jean-Michel Vernes Y. Gloria Meng Samantha Lein Pamela Chan Jed Ross Richard Carano Rong Deng Nicholas Lewin-Koh Min Xu Becket Feierbach 《PLoS pathogens》2014,10(4)
Human cytomegalovirus (HCMV) is the most common cause of congenital virus infection. Congenital HCMV infection occurs in 0.2–1% of all births, and causes birth defects and developmental abnormalities, including sensorineural hearing loss and developmental delay. Several key studies have established the guinea pig as a tractable model for the study of congenital HCMV infection and have shown that polyclonal antibodies can be protective [1]–[3]. In this study, we demonstrate that an anti-guinea pig CMV (GPCMV) glycoprotein H/glycoprotein L neutralizing monoclonal antibody protects against fetal infection and loss in the guinea pig. Furthermore, we have delineated the kinetics of GPCMV congenital infection, from maternal infection (salivary glands, seroconversion, placenta) to fetal infection (fetus and amniotic fluid). Our studies support the hypothesis that a neutralizing monoclonal antibody targeting an envelope GPCMV glycoprotein can protect the fetus from infection and may shed light on the therapeutic intervention of HCMV congenital infection in humans. 相似文献
993.
Joseph A Hampel Samantha N Spath Ingrid L Bergin Ailam Lim Steven R Bolin Melissa C Dyson 《Comparative medicine》2014,64(6):478-485
Hemotrophic mycoplasma (hemoplasma) infection in research sheep can confound experimental results and contribute to morbidity and mortality. Prevalence and clinicopathologic studies historically relied on blood-smear diagnosis, but systematic studies using current molecular techniques are warranted. Here we sought to report the prevalence of subclinical infection in our study population, compare diagnostic sensitivity and specificity between blood smears and a PCR assay, and determine the effects of infection on CBC variables and erythrocyte membrane fragility. We collected whole-blood samples from 111 convenience-sampled research sheep. All samples were tested for hemoplasmas by using a PCR assay, blood smears were evaluated for visual presence of hemoplasmas, and CBC and osmotic fragility assays were performed. Subclinical prevalence, according to PCR diagnosis, was 14.1% (14 of 99) in our study population. Relative to the PCR assay, blood-smear diagnosis was 8.3% sensitive and 100% specific for hemoplasma detection. Subclinical infection was associated with changes in MCV, MCHC, RBC distribution width, and absolute monocyte count. Acute infection was associated with changes in RBC mass, Hgb concentration, MCV, MCH, MCHC, and absolute lymphocyte and monocyte counts. Acute infection was associated with increased mean erythrocyte fragility compared with that in uninfected control and treated sheep. We demonstrated that hemoplasma infection is common in our study population, blood-smear evaluation is insensitive at detecting infection, and infection is associated with changes in CBC variables and increased erythrocyte membrane fragility. These findings raise concerns regarding the suitability of hemoplasma-infected sheep for biomedical research.Abbreviations: MEF, mean erythrocyte fragilityHemoplasma infection in sheep is caused by Mycoplasma ovis and ‘Candidatus M. haemovis.’15,34 Recent work, however, suggests M. ovis and ‘Ca. Mycoplasma haemovis’ represent the same organism with 2 different copies of the 16S rRNA.11 This agent, formerly called Eperythrozoon ovis, is a cell-wall–free bacterial parasite that is intimately associated with the plasma membrane of sheep erythrocytes.25 It typically is considered to be nonpathogenic in chronic infection, but it occasionally is associated with hemolytic anemia during acute infection.24 Hemoplasma infection can confound experimental results and contribute to morbidity and mortality in research animals.20 Infections have been reported worldwide. The clinicopathologic effects in lab animals may be a concern for biomedical research.2The organism cannot be grown in culture, making its diagnosis difficult. Previous prevalence and clinicopathologic studies relied on blood-smear diagnosis or serology.5,6,8-10,13,14,16,20,22,25-27,31 Systematic studies on the diagnosis and clinicopathologic effects of ovine hemoplasma infection using current, molecular techniques are unavailable. In addition, the contemporary prevalence of hemoplasma infection in research sheep in the United States is unknown.The purpose of the current study was to evaluate: 1) the prevalence of subclinical hemoplasma infection in our study population of research sheep; 2) the sensitivity and specificity of blood smears to detect hemoplasma infection; 3) the effects of subclinical and acute hemoplasma infection on CBC variables; and 4) the effects of acute hemoplasma infection on erythrocyte membrane fragility. We hypothesized that: 1) subclinical hemoplasma infection is common; 2) the examination of blood smears is not as sensitive or specific as is PCR analysis for detecting hemoplasma infection; 3) subclinical and acute infection alters CBC variables; and 4) acute infection increases erythrocyte membrane fragility.To address these questions, we collected whole-blood samples from 111 convenience-sampled research sheep as part of routine health surveillance. All samples were PCR tested by using hemoplasma-specific primers, blood smears were evaluated, and CBC analyses were performed. Osmotic fragility assays were performed on a subset of animals. 相似文献
994.
The second committed step in chlorophyll biosynthesis is the transfer of a methyl group from S-adenosyl-l-methionine (SAM) to magnesium protoporphyrin IX (MgP) forming MgP monomethylester (MgPME). This reaction is catalyzed by the enzyme MgP methyltransferase (ChlM). Previous investigation of this enzyme has involved the use of time-consuming techniques requiring separation of products from substrates. More recent methyltransferase studies use coupling enzymes to monitor changes in absorption/fluorescence for the measurement of activity. However, due to the spectral properties of porphyrins, many of these assays are unsuitable for analysis of the catalytic properties of ChlM. Here we report the successful development of a coupled, continuous spectrophotometric assay to measure the activity of ChlM. The product of the methyltransferase reaction, S-adenosyl-l-homocysteine (SAH), is converted into adenine and then hypoxanthine by the recombinant coupling enzymes SAH nucleosidase and adenine deaminase, respectively. The appearance of hypoxanthine results in a decrease in absorbance at 265 nm.The utility of this assay was shown by the characterization of ChlM from the cyanobacterium Synechocystis sp. PCC 6803. Kinetic parameters obtained support data acquired using the discontinuous HPLC-based assay and provide further evidence for the stimulation of ChlM by the H subunit of magnesium chelatase (ChlH). 相似文献
995.
Lipid droplets (LDs) are key cellular organelles involved in lipid storage and mobilisation. While the major signalling cascades and many of the regulators of lipolysis have been identified, the cellular interactions involved in lipid mobilisation and release remain largely undefined. In non-adipocytes, LDs are small, mobile and interact with other cellular compartments. In contrast, adipocytes primarily contain very large, immotile LDs. The striking morphological differences between LDs in adipocytes and non-adipocytes suggest that key differences must exist in the manner in which LDs in different cell types interact with other organelles. Recent studies have highlighted the complexity of LD interactions, which can be both homotypic, with each other, and heterotypic, with other organelles. The molecules involved in these interactions are also now emerging, including Rab proteins, key regulators of membrane traffic, and caveolin, an integral membrane protein providing a functional link between the cell surface and LDs. Here we summarise recent insights into the cell biology of the LD particularly focussing on the homotypic and heterotypic interactions in both adipocytes and non-adipocytes. We speculate that these interactions may involve inter-organelle membrane contact sites or a hemi-fusion type mechanism to facilitate lipid transfer. 相似文献
996.
997.
Xiao-Yan Du Brian A. Zabel Timothy Myles Samantha J. Allen Tracy M. Handel Peter P. Lee Eugene C. Butcher Lawrence L. Leung 《The Journal of biological chemistry》2009,284(2):751-758
Chemerin is a potent chemoattractant for cells expressing the serpentine
receptor CMKLR1 (chemokine-like receptor 1), such as plasmacytoid dendritic
cells and tissue macrophages. The bioactivity of chemerin is
post-translationally regulated; the attractant circulates in blood in a
relatively inactive form (prochemerin) and is activated by carboxyl-terminal
proteolytic cleavage. We discovered that plasma carboxypeptidase N (CPN) and B
(CPB or activated thrombin-activable fibrinolysis inhibitor, TAFIa) enhanced
the bioactivity of 10-mer chemerin peptide NH2-YFPGQFAFSK-COOH by
removing the carboxyl-terminal lysine (K). Sequential cleavages of either a
prochemerin peptide (NH2-YFPGQFAFSKALPRS-COOH) or recombinant
full-length prochemerin by plasmin and CPN/CPB substantially increased their
chemotactic activities. Endogenous CPN present in circulating plasma enhanced
the activity of plasmin-cleaved prochemerin. In addition, we discovered that
platelets store chemerin protein and release it upon stimulation. Thus
circulating CPN/CPB and platelets may potentially contribute to regulating the
bioactivity of leukocyte chemoattractant chemerin, and further extend the
molecular link between blood coagulation/fibrinolysis and CMKLR1-mediated
immune responses.Chemerin is a recently discovered chemoattractant molecule that is
predicted to share structural similarity with cystatins (cysteine protease
inhibitors) and cathelicidin precursors (antibacterial peptides)
(1). Chemerin is present in
circulating blood and several human inflammatory fluids
(1). Even though chemerin is
not similar to CXC and CC chemokines based on primary amino acid sequence, it
functions like a chemokine in that it induces leukocyte migration and
intracellular calcium mobilization. Chemerin receptor chemokine-like receptor
1 (CMKLR1,3 also named
ChemR23) is a G protein-coupled receptor specifically expressed by circulating
human plasmacytoid dendritic cells, natural killer cells, and tissue
macrophages
(1–5).
In their capacity as antigen-presenting cells, plasmacytoid dendritic cells
and macrophages can influence the activation of many other cell types,
including monocytes, myeloid dendritic cells, B cells, T cells, and natural
killer cells; thus chemerin appears to be an important chemoattractant in both
innate and adaptive immune responses
(2,
6,
7).Chemerin circulates in blood in an inactive prochemerin form at low
nanomolar concentrations (∼3 nm)
(4). Its chemotactic activity
is released following proteolytic cleavage of its carboxyl-terminal amino
acids by serine proteases of the coagulation, fibrinolytic, and inflammatory
cascades (4,
8). These include factor XIIa,
VIIa, plasmin, neutrophil elastase, and mast cell tryptase. Of interest,
staphopain B, a cysteine protease secreted by Staphylococcus aureus,
also cleaves prochemerin and converts it into a potent chemoattractant
(9). Interestingly, the
cleavage sites in the labile carboxyl terminus
(NH2-YFPGQFAFSKALPRS-COOH) are not conserved, and the cleavage
products generated by chemerin-activating proteases display different
potencies in bioactivity assays. Based on synthetic peptides, the 9-mer
NH2-YFPGQFAFS-COOH is the most active, but it is still not as
active as intact cleaved chemerin protein, indicating that the amino-terminal
part of chemerin is required for maximal activity
(4,
10).Plasma carboxypeptidases CPN and CPB cleave the basic amino acids arginine
or lysine from the carboxyl terminus of proteins or peptides such as
bradykinin and complement proteins C3a and C5a. CPN is a constitutively active
zinc metalloprotease present in plasma at a concentration of about 100
nm and is considered the major anaphylatoxins inhibitor
(11), generating inactive
“desArg” forms of C3a and C5a. In contrast, CPB exists in plasma
as a proenzyme, proCPB, or thrombin-activable fibrinolysis inhibitor (TAFI) at
a concentration of about 50 nm and is activated by thrombin in
complex with thrombomodulin on the vascular endothelial surface. CPB inhibits
fibrin degradation by removing carboxyl-terminal lysines from partially
digested fibrin, which prevents further incorporation of fibrinolytic
plasminogen and tissue plasminogen activator
(12,
13). CPB is thermolabile and
has a half-life of ∼15 min at 37 °C
(14). We have shown that CPB
also has broad substrate reactivity and is able to cleave and inactivate
bradykinin, C3a, C5a, and thrombin-cleaved osteopontin
(15–17).
CPN and CPB may play complementary roles, with the former being constitutively
active and capable of regulating systemic anaphylatoxins, and the latter
activated locally at sites of vascular injury to provide site-specific
anti-inflammatory control. Peptidases can also modulate the biological
activity of certain chemokines
(4). For example, dipeptidyl
peptidase (DPP-IV/CD26), a serine protease, inactivates CXCL9, CXCL10, CXCL11,
and CXCL12 by cleaving these chemokines in the amino terminus
(18,
19).Platelets store a variety of potent cytokines and chemokines within
α-granules that are released upon cell activation. Platelet
degranulation products, particularly the leukocyte chemoattractants, which
include CXCL4 (platelet factor 4), β-thromboglobulin, CCL5 (RANTES), CCL7
(monocyte chemotactic protein 3), and CXCL12 (stromal-derived factor 1), may
contribute to host defense and also play a role in pathophysiologic conditions
(20,
21). For example, platelet
factor 4 forms complexes with heparin in blood or some glycosaminoglycans on
platelet surfaces to form the major antigen implicated in heparin-induced
thrombocytopenia (22,
23). Platelets not only store
CXCL12 but also express its receptor CXCR4, a coreceptor for cellular entry of
human immunodeficiency virus, type 1, suggesting that platelets may be
involved in host defense
(24).In this study, we found that plasma CPN or CPB can function in concert with
plasmin to elicit and augment the chemotactic activity of prochemerin.
Furthermore, we show that platelets could store and release partially active
chemerin upon activation. Thus circulating CPN/CPB and platelets may
contribute to regulating the bioactivity of leukocyte chemoattractant chemerin
and further extend the molecular link between blood coagulation/fibrinolysis
and CMKLR1-mediated immune responses. 相似文献
998.
Fang Cao Xiangzhi Li Samantha Hiew Hugh Brady Yifan Liu Yali Dou 《RNA (New York, N.Y.)》2009,15(7):1274-1281
Small RNAs play important roles in the establishment and maintenance of heterochromatin structures. We show the presence of telomere specific small RNAs (tel-sRNAs) in mouse embryonic stem cells that are ∼24 nucleotides in length, Dicer-independent, and 2′-O-methylated at the 3′ terminus. The tel-sRNAs are asymmetric with specificity toward telomere G-rich strand, and evolutionarily conserved from protozoan to mammalian cells. Furthermore, tel-sRNAs are up-regulated in cells that carry null mutation of H3K4 methyltransferase MLL (Mll(−/−)) and down-regulated in cells that carry null mutations of histone H3K9 methyltransferase SUV39H (Suv39h1/h2(−/−)), suggesting that they are subject to epigenetic regulation. These results support that tel-sRNAs are heterochromatin associated pi-like small RNAs. 相似文献
999.
1000.
Distance software: design and analysis of distance sampling surveys for estimating population size 总被引:2,自引:0,他引:2