全文获取类型
收费全文 | 341篇 |
免费 | 18篇 |
国内免费 | 9篇 |
专业分类
368篇 |
出版年
2023年 | 2篇 |
2022年 | 4篇 |
2021年 | 2篇 |
2016年 | 2篇 |
2015年 | 6篇 |
2014年 | 9篇 |
2013年 | 15篇 |
2012年 | 10篇 |
2011年 | 28篇 |
2010年 | 8篇 |
2009年 | 6篇 |
2008年 | 11篇 |
2007年 | 15篇 |
2006年 | 10篇 |
2005年 | 10篇 |
2004年 | 11篇 |
2003年 | 9篇 |
2002年 | 4篇 |
2001年 | 11篇 |
2000年 | 9篇 |
1999年 | 5篇 |
1998年 | 5篇 |
1997年 | 2篇 |
1995年 | 7篇 |
1993年 | 2篇 |
1992年 | 8篇 |
1991年 | 14篇 |
1990年 | 7篇 |
1989年 | 9篇 |
1988年 | 10篇 |
1987年 | 15篇 |
1986年 | 11篇 |
1985年 | 13篇 |
1984年 | 8篇 |
1983年 | 3篇 |
1982年 | 6篇 |
1981年 | 13篇 |
1980年 | 7篇 |
1979年 | 5篇 |
1978年 | 7篇 |
1977年 | 6篇 |
1976年 | 4篇 |
1974年 | 2篇 |
1970年 | 3篇 |
1965年 | 1篇 |
1964年 | 1篇 |
1962年 | 1篇 |
1957年 | 3篇 |
1955年 | 1篇 |
1953年 | 1篇 |
排序方式: 共有368条查询结果,搜索用时 15 毫秒
91.
P Manjunath Y L Marcel J Uma N G Seidah M Chrétien A Chapdelaine 《The Journal of biological chemistry》1989,264(28):16853-16857
Bovine seminal plasma contains four similar acidic proteins, previously designated as BSP (bovine seminal plasma)-A1, BSP-A2, BSP-A3, and BSP-30-kDa, that when added to pituitary cell cultures result in the immediate secretion of gonadotropins (follitropin and lutropin). However, when calf or horse serum was included in the culture medium the secretion of gonadotropins was completely prevented. This effect was seen at levels up to 200 micrograms of BSP protein/ml while the presence of more than 200 micrograms of BSP protein/ml in the serum medium continued to release gonadotropins. This could be explained by the presence in the sera of a binding factor to the BSP proteins which prevents their action. This binding factor has been detected in all the sera tested, including human serum, in dot-blot experiments using 125I-labeled BSP-A1, -A2, -A3, or -30-kDa protein. Thus, it was of interest to isolate this binding factor from human serum by affinity chromatography on a column of BSP-A1/-A2-agarose. The purified binding factor was then identified as apolipoprotein A-I (apoA-I) by the following criteria: (a) it has a molecular mass of 27,000 daltons, (b) the amino acid composition is similar to apoA-I, (c) the first 25 residues at the amino-terminal end of this binding factor are identical to apoA-I, and (d) the binding factor cross-reacts in the radioimmunoassay of apoA-I. Furthermore, BSP proteins also bind to purified plasma apoA-I and apoA-I associated with high density lipoprotein. ApoA-I is the major protein of plasma high density lipoprotein and plays an important role in lipid transport and metabolism. Thus, the binding of bovine seminal plasma proteins to apoA-I suggests some physiological significance in lipoprotein function or vice versa. 相似文献
92.
目的:尝试应用RNA干扰(RNAi)沉默猪源PK-15细胞中的猪内源性反转录病毒(PERV),并通过反转录酶活性及pol基因相对荧光定量PCR检测沉默效果。方法:依据GenBank公布的PERV pol基因序列,采用Invitro-gen公司的BLOCK-iT RNAi Designer软件设计Stealth小干扰RNA(siRNA)序列;将合成的siRNA转染PK-15细胞,72 h后检测细胞上清PERV反转录酶活性及细胞内pol基因拷贝数并评价沉默效果。结果:反转录酶活性及pol基因拷贝数检测结果表明,设计的3条Stealth siRNA序列中,位于pol基因3272~3296 bp的序列能有效沉默PERV。结论:RNAi方法可有效使猪源PK-15细胞中的PERV沉默,为进一步研究天然抗病毒分子与PERV的相互作用提供了实验基础,同时也为猪源异种移植研究中去除PERV提供了一种可供尝试的方法。 相似文献
93.
缺血性脑卒中患者同型半胱氨酸代谢相关酶基因突变的研究 总被引:4,自引:0,他引:4
为研究同型半胱氨酸代谢相关酶亚甲基四氢叶酸还原酶(MTHFR)基因C677T和胱硫醚-β-合成酶(CBS)基因T833C位点碱基突变与缺血性脑卒中的关系,对74例缺血性脑卒中患者和83例健康对照者,采用聚合酶链反应-限制性片段长度多态性(PCR-RFLP)技术检测MTHFR基因C677T基因型,用扩增阻滞突变体系法(ARMS)检测CBS基因T833C突变。实验检出患者组MTHFR基因T纯合基因型、杂合基因型和T等位基因频率分别为2.7%、51.4%和28.4%,对照组分别为1.2%、39,8%和21.1%。患者组CBS基因C纯合基因型和C等位基因频率分别为13.5%和43.9%,对照组分别为6.0%和38.0%。Multiple Logistic Regression分析显示;C677T位点T等位基因,T833C位点C等位基因以及年龄均与缺血性脑卒中发病有关(P<0.05),C677T位点T等位基因的比值比(OR)为1.74(95%CI 1.06~2,B6)和T833C位点C等位基因的比值比为1.73(95%CI 1.07~-2.81)。实验显示MTHFR C677T和CBS T833C基因位点突变与缺血性脑卒中发病有关,上述两个基因位点突变可能是缺血性脑卒中发病的遗传因素。 相似文献
94.
短尾猴和日本猴雄性性行为的比较研究 总被引:1,自引:2,他引:1
本文比较研究了短尾猴和日本猴的雄性性行为。短尾猴属单次爬跨射精型种类,每次交配平均持续23.2秒,日本猴属多次爬跨射精型种类,每次交配平均持续8.2分,交配期间雄性平均爬跨雌性lO.6次才能达到射精。短尾猴第1顺位雄性是群中的主要交配者,它占有交配总数的70.9 %;而日本猴第5顺位以下的雄性占交配总数的64%。交配中,两种猴雄性间相互打搅行为的发生频率大致相当。短尾猴高顺位雄性的打搅行为能使低顺位雄性的交配中断,但日本猴高顺位的打搅行为只能使低顺位雄性交配的54.3%中断。 相似文献
95.
A Basak Y T Gong J A Cromlish J A Paquin F Jean N G Seidah C Lazure M Chrétien 《International journal of peptide and protein research》1990,36(1):7-17
Eight argininal semicarbazone containing peptides prepared by liquid phase synthesis were all found to be reversible inhibitors of model serine proteinases including trypsin and plasma kallikrein (PK). Among the peptides tested, those having a Lys residue at position P2 displayed the maximum binding potency towards PK. One of the peptides, Leu-enkephalin-argininal semicarbazone, a comparatively weak inhibitor, was chosen in order to develop an affinity-based purification protocol for PK. The affinity column was prepared by covalent attachment of the NH2-terminal moiety of the peptidyl semicarbazone to a solid-phase matrix bearing a spacer group. For efficient binding of PK, it was found necessary to optimize parameters like the concentration of inhibitor linked to the solid matrix, the ionic strength of the buffer used, the temperature and the pH. The majority of the bound enzyme could be recovered following elution with guanidine hydrochloride or benzamidine hydrochloride in a high salt buffer at pH 6.0. The usefulness of the affinity procedure towards the purification of other serine proteinases is also discussed. 相似文献
96.
A crucial step in the life cycle of arenaviruses is the biosynthesis of the mature fusion-active viral envelope glycoprotein (GP) that is essential for virus-host cell attachment and entry. The maturation of the arenavirus GP precursor (GPC) critically depends on proteolytic processing by the cellular proprotein convertase (PC) subtilisin kexin isozyme-1 (SKI-1)/site-1 protease (S1P). Here we undertook a molecular characterization of the SKI-1/S1P processing of the GPCs of the prototypic arenavirus lymphocytic choriomeningitis virus (LCMV) and the pathogenic Lassa virus (LASV). Previous studies showed that the GPC of LASV undergoes processing in the endoplasmic reticulum (ER)/cis-Golgi compartment, whereas the LCMV GPC is cleaved in a late Golgi compartment. Herein we confirm these findings and provide evidence that the SKI-1/S1P recognition site RRLL, present in the SKI-1/S1P prodomain and LASV GPC, but not in the LCMV GPC, is crucial for the processing of the LASV GPC in the ER/cis-Golgi compartment. Our structure-function analysis revealed that the cleavage of arenavirus GPCs, but not cellular substrates, critically depends on the autoprocessing of SKI-1/S1P, suggesting differences in the processing of cellular and viral substrates. Deletion mutagenesis showed that the transmembrane and intracellular domains of SKI-1/S1P are dispensable for arenavirus GPC processing. The expression of a soluble form of the protease in SKI-I/S1P-deficient cells resulted in the efficient processing of arenavirus GPCs and rescued productive virus infection. However, exogenous soluble SKI-1/S1P was unable to process LCMV and LASV GPCs displayed at the surface of SKI-I/S1P-deficient cells, indicating that GPC processing occurs in an intracellular compartment. In sum, our study reveals important differences in the SKI-1/S1P processing of viral and cellular substrates. 相似文献
97.
Kim W Essalmani R Szumska D Creemers JW Roebroek AJ D'Orleans-Juste P Bhattacharya S Seidah NG Prat A 《Molecular and cellular biology》2012,32(17):3382-3391
In mammals, seven proprotein convertases (PCs) cleave secretory proteins after basic residues, and four of them are called furin-like PCs: furin, PC5, PACE4, and PC7. In vitro, they share many substrates. However, furin is essential during development since deficient embryos die at embryonic day 11 and exhibit multiple developmental defects, particularly defects related to the function of endothelial cells. To define the role of furin in endothelial cells, an endothelial cell-specific knockout (ecKO) of the Furin gene was generated. Newborns die shortly after birth, indicating that furin is essential in these cells. Magnetic resonance imaging revealed that ecKO embryos exhibit ventricular septal defects (VSD) and/or valve malformations. In addition, primary cultures of wild-type and ecKO lung endothelial cells revealed that ecKO cells are unable to grow. Growth was efficiently rescued by extracellular soluble furin. Analysis of the processing of precursors of endothelin-1 (ET-1), adrenomedullin (Adm), transforming growth factor β1 (TGF-β1), and bone morphogenetic protein 4 (BMP4) confirmed that ET-1, Adm, and TGF-β1 are in vivo substrates of endothelial furin. Mature ET-1 and BMP4 forms were reduced by ~90% in ecKO purified endothelial cells from lungs. 相似文献
98.
99.
Kulandaivelu S. Vetrivel Xavier Meckler Ying Chen Phuong D. Nguyen Nabil G. Seidah Robert Vassar Philip C. Wong Masaki Fukata Maria Z. Kounnas Gopal Thinakaran 《The Journal of biological chemistry》2009,284(6):3793-3803
Alzheimer disease β-amyloid (Aβ) peptides are generated via
sequential proteolysis of amyloid precursor protein (APP) by BACE1 and
γ-secretase. A subset of BACE1 localizes to cholesterol-rich membrane
microdomains, termed lipid rafts. BACE1 processing in raft microdomains of
cultured cells and neurons was characterized in previous studies by disrupting
the integrity of lipid rafts by cholesterol depletion. These studies found
either inhibition or elevation of Aβ production depending on the extent
of cholesterol depletion, generating controversy. The intricate interplay
between cholesterol levels, APP trafficking, and BACE1 processing is not
clearly understood because cholesterol depletion has pleiotropic effects on
Golgi morphology, vesicular trafficking, and membrane bulk fluidity. In this
study, we used an alternate strategy to explore the function of BACE1 in
membrane microdomains without altering the cellular cholesterol level. We
demonstrate that BACE1 undergoes S-palmitoylation at four Cys
residues at the junction of transmembrane and cytosolic domains, and Ala
substitution at these four residues is sufficient to displace BACE1 from lipid
rafts. Analysis of wild type and mutant BACE1 expressed in BACE1 null
fibroblasts and neuroblastoma cells revealed that S-palmitoylation
neither contributes to protein stability nor subcellular localization of
BACE1. Surprisingly, non-raft localization of palmitoylation-deficient BACE1
did not have discernible influence on BACE1 processing of APP or secretion of
Aβ. These results indicate that post-translational
S-palmitoylation of BACE1 is not required for APP processing, and
that BACE1 can efficiently cleave APP in both raft and non-raft
microdomains.Alzheimer disease-associated β-amyloid
(Aβ)3 peptides
are derived from the sequential proteolysis of β-amyloid precursor
protein (APP) by β- and γ-secretases. The major β-secretase is
an aspartyl protease, termed BACE1 (β-site
APP-cleaving enzyme 1)
(1–4).
BACE1 cleaves APP within the extracellular domain of APP, generating the N
terminus of Aβ. In addition, BACE1 also cleaves to a lesser extent within
the Aβ domain between Tyr10 and Glu11
(β′-cleavage site). Processing of APP at these sites results in the
shedding/secretion of the large ectodomain (sAPPβ) and generating
membrane-tethered C-terminal fragments +1 and +11 (β-CTF)
(5). The multimeric
γ-secretase cleaves at multiple sites within the transmembrane domain of
β-CTF, generating C-terminal heterogeneous Aβ peptides (ranging in
length between 38 and 43 residues) that are secreted, as well as cytosolic APP
intracellular domains (6). In
addition to BACE1, APP can be cleaved by α-secretase within the Aβ
domain between Lys16 and Leu17, releasing sAPPα
and generating α-CTF. γ-Secretase cleavage of α-CTF
generates N-terminal truncated Aβ, termed p3.Genetic ablation of BACE1 completely abolishes Aβ production,
establishing BACE1 as the major neuronal enzyme responsible for initiating
amyloidogenic processing of APP
(4,
7). Interestingly, both the
expression and activity of BACE1 is specifically elevated in neurons adjacent
to senile plaques in brains of individuals with Alzheimer disease
(8). In the past few years
additional substrates of BACE1 have been identified that include APP
homologues APLP1 and APLP2 (9),
P-selectin glycoprotein ligand-1
(10), β-galactoside
α2,6-sialyltransferase
(11), low-density lipoprotein
receptor-related protein (12),
β-subunits of voltage-gated sodium channels
(13), and neuregulin-1
(14,
15), thus extending the
physiological function of BACE1 beyond Alzheimer disease pathogenesis.BACE1 is a type I transmembrane protein with a long extracellular domain
harboring a catalytic domain and a short cytoplasmic tail. BACE1 is
synthesized as a proenzyme, which undergoes post-translational modifications
that include removal of a pro-domain by a furin-like protease,
N-glycosylation, phosphorylation, S-palmitoylation, and
acetylation, during the transit in the secretory pathway
(16–20).
In non-neuronal cells the majority of BACE1 localizes to late Golgi/TGN and
endosomes at steady-state and a fraction of BACE1 also cycles between the cell
surface and endosomes (21).
The steady-state localization of BACE1 is consistent with the acidic pH
optimum of BACE1 in vitro, and BACE1 cleavage of APP is observed in
the Golgi apparatus, TGN, and endosomes
(22–25).
BACE1 endocytosis and recycling are mediated by the GGA family of adaptors
binding to a dileucine motif (496DISLL) in its cytoplasmic tail
(21,
26–31).
Phosphorylation at Ser498 within this motif modulates GGA-dependent
retrograde transport of BACE1 from endosomes to TGN
(21,
26–31).Over the years, a functional relationship between cellular cholesterol
level and Aβ production has been uncovered, raising the intriguing
possibility that cholesterol levels may determine the balance between
amyloidogenic and non-amyloidogenic processing of APP
(32–34).
Furthermore, several lines of evidence from in vitro and in
vivo studies indicate that cholesterol- and sphingolipid-rich membrane
microdomains, termed lipid rafts, might be the critical link between
cholesterol levels and amyloidogenic processing of APP. Lipid rafts function
in the trafficking of proteins in the secretory and endocytic pathways in
epithelial cells and neurons, and participate in a number of important
biological functions (35).
BACE1 undergoes S-palmitoylation
(19), a reversible
post-translational modification responsible for targeting a variety of
peripheral and integral membrane proteins to lipid rafts
(36). Indeed, a significant
fraction of BACE1 is localized in lipid raft microdomains in a
cholesterol-dependent manner, and addition of glycosylphosphatidylinositol
(GPI) anchor to target BACE1 exclusively to lipid rafts increases APP
processing at the β-cleavage site
(37,
38). Antibody-mediated
co-patching of cell surface APP and BACE1 has provided further evidence for
BACE1 processing of APP in raft microdomains
(33,
39). Components of the
γ-secretase complex also associate with detergent-resistant membrane
(DRM) fractions enriched in raft markers such as caveolin, flotillin, PrP, and
ganglioside GM1 (40). The
above findings suggest a model whereby APP is sequentially processed by BACE1
and γ-secretase in lipid rafts.Despite the accumulating evidence, cleavage of APP by BACE1 in non-raft
membrane regions cannot be unambiguously ruled out because of the paucity of
full-length APP (APP FL) and BACE1 in DRM isolated from adult brain and
cultured cells (41). Moreover,
it was recently reported that moderate reduction of cholesterol (<25%)
displaces BACE1 from raft domains, and increases BACE1 processing by promoting
the membrane proximity of BACE1 and APP in non-raft domains
(34). Nevertheless, this study
also found that BACE1 processing of APP is inhibited with further loss of
cholesterol (>35%), consistent with earlier studies
(32,
33). Nevertheless, given the
pleiotropic effects of cholesterol depletion on membrane properties and
vesicular trafficking of secretory and endocytic proteins
(42–47),
unequivocal conclusions regarding BACE1 processing of APP in lipid rafts
cannot be reached based on cholesterol depletion studies.In this study, we explored the function of BACE1 in lipid raft microdomains
without manipulating cellular cholesterol levels. In addition to the
previously reported S-palmitoylation sites
(Cys478/Cys482/Cys485) within the cytosolic
tail of BACE1 (19), we have
identified a fourth site (Cys474) within the transmembrane domain
of BACE1 that undergoes S-palmitoylation. A BACE1 mutant with Ala
substitution of all four Cys residues (BACE1-4C/A) fails to associate with DRM
in cultured cells, but is not otherwise different from wtBACE1 in terms of
protein stability, maturation, or subcellular localization. Surprisingly, APP
processing and Aβ generation were unaffected in cells stably expressing
the BACE1-4C/A mutant. Finally, we observed an increase in the levels of APP
CTFs in detergent-soluble fractions of BACE1-4C/A as compared with wtBACE1
cells. Thus, our data collectively indicate a non-obligatory role of
S-palmitoylation and lipid raft localization of BACE1 in
amyloidogenic processing of APP. 相似文献
100.