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1.
Characterization of ovarian gonadotropin receptor. Monomer and associated form of the receptor 总被引:3,自引:0,他引:3
We have purified luteinizing hormone/human choriogonadotropin (hCG) receptor from rat ovary by sequential affinity column on wheat germ lectin-Sepharose and hCG-Sepharose chromatography. The purified receptor, previously identified as a single protein on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) (Kusuda, S., and Dufau, M.L. (1986) J. Biol. Chem. 261, 16161-16168), was further characterized by radioiodination with 1,3,4,6-tetrachloro-3 alpha, 6 alpha-diphenylglycouril, and column chromatography on wheat germ lectin-Sepharose. Autoradiography of SDS-PAGE analysis under reducing conditions showed a single radiolabeled band of Mr = 80,000. The radioiodinated receptors treated with peptide:N-glycosidase F migrated at Mr = 54,000. Treatment with neuraminidase alone caused only a minor reduction in molecular weight, and subsequent treatment with endo-alpha-N-acetyl-D-galactosaminidase had little further effect on the receptor. When the radioiodinated receptor was analyzed by fast protein liquid chromatography, a single broad peak was eluted with Mr of approximately 350,000. The higher Mr of radioiodinated receptors than that of native receptors (Mr = 190,000 dimeric form) could be due to the aggregation of labeled molecules. These complexes dissociated into the monomeric form in the presence of SDS. To determine whether the monomers can bind hormone, the purified unlabeled receptors resolved with SDS were electroblotted to nitrocellulose membranes and incubated with 125I-hCG. Autoradiograms of the blots showed a band of monomer (Mr = 78,000) as well as one of dimer (Mr approximately 150,000). These studies have demonstrated that the luteinizing hormone/hCG receptors are predominantly N-linked glycosylated and suggest that the native receptor is a dimer of identical hormone binding subunits associated by noncovalent interactions. Although the individual subunits can bind hormone, it is conceivable that the dimeric form is necessary for signal transduction. 相似文献
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Junko Komura Ikumi Tamai Mizuho Senmaru Tetsuya Terasaki Yoshimichi Sai Akira Tsuji 《Journal of neurochemistry》1996,67(1):330-335
Abstract: The characteristics of β-alanine transport at the blood-brain barrier were studied by using primary cultured bovine brain capillary endothelial cells. Kinetic analysis of the β-[3 H]alanine transport indicated that the transporter for β-alanine functions with Kt of 25.3 ± 2.5 µ M and J max of 6.90 ± 0.48 nmol/30 min/mg of protein in the brain capillary endothelial cells. β-[3 H]Alanine uptake is mediated by an active transporter, because metabolic inhibitors (2,4-dinitrophenol and NaN3 ) and low temperature reduced the uptake significantly. Furthermore, the uptake of β-[3 H]alanine required Na+ and Cl− in the external medium. Stoichiometric analysis of the transport demonstrated that two sodium ions and one chloride ion are associated with one β-alanine molecule. The Na+ and Cl− -dependent uptake of β-[3 H]alanine was stimulated by a valinomycin-induced inside-negative K+ -diffusion potential. β-Amino acids (β-alanine, taurine, and hypotaurine) inhibited strongly the uptake of β-[3 H]alanine, whereas α- and γ-amino acids had little or no inhibitory effect. In ATP-depleted cells, the uptake of β-[3 H]alanine was stimulated by preloading of β-alanine or taurine but not l -leucine. These results show that β-alanine is taken up by brain capillary endothelial cells, via the secondary active transport mechanism that is common to β-amino acids. 相似文献
4.
Mizuho Komatsu-Takaki 《FEBS letters》1984,175(2):433-438
Decay of light-triggered ATP hydrolysis in the dark was diminished with a decrease in chloroplast concentration. The enhancing effect of NH4Cl on ATP hydrolysis decreased with dark time. The decrease was much faster than that in ATP hydrolysis activity. The NH4Cl effect increased with ATP preincubation time. Reactivation of ATP hydrolysis occurred with the progress of ATP hydrolysis. Pi enhanced the activation remarkably. These results suggest that ATP hydrolysis produces some energized state, which stimulates NH4C1 effect and makes coupling factor active in the presence of Pi and that to keep coupling factor active, energy is not necessarily needed. 相似文献
5.
Kouichi Yasuda Yoko Nakayama Mizuho Tanaka Mikiko Tanaka Ryota Mori Kiyofumi Furusawa 《Somatosensory & motor research》2013,30(1):30-35
The present study was an attempt to identify the location of genioglossal respiratory and swallowing motoneuron cell bodies within the hypoglossal (XII) nucleus using both electrophysiological and morphological studies. The genioglossus muscle is innervated by the genioglossal branch of the medial XII nerve. At the entrance to the muscle, the genioglossal branch divides in the directions of the mandible and tongue. Five of five rats displayed both respiratory-related and swallowing-related bursts in the medial XII branch towards the mandible. All five rats also displayed swallowing-related bursts in the medial XII branch towards the tongue. In addition, horseradish peroxidase conjugated to wheatgerm agglutinin (HRP:WGA) was injected into the proximal cut ends of each branch. When HRP:WGA was injected into the branch in the direction of the mandible, HRP-labeled cells were detected in the lateral region of the ventromedial subnucleus in the XII nucleus, extending from 0.7 to 1.2 mm rostral to the obex. On the other hand, after injection into the branch in the direction of the mandible, HRP-labeled cells were detected in the ventromedial subnucleus of the XII nucleus, extending from 0.3 to 1.2 mm rostral to the obex. These results provide evidence that genioglossal respiration-related and swallowing-related motoneurons are located in different portions within the ventromedial subnucleus of the XII nucleus. 相似文献
6.
Naoko Sekino‐Suzuki Kohei Yuyama Toshiaki Miki Mizuho Kaneda Hidenori Suzuki Naomasa Yamamoto Tadashi Yamamoto Chitose Oneyama Masato Okada Kohji Kasahara 《Journal of neurochemistry》2013,124(4):514-522
The association of gangliosides with specific proteins in the central nervous system was examined by coimmunoprecipitation with an anti‐ganglioside antibody. The monoclonal antibody to the ganglioside GD3 (R24) immunoprecipitated the Csk (C‐terminal src kinase)‐binding protein (Cbp). Sucrose density gradient analysis showed that Cbp of rat cerebellum was detected in detergent‐resistant membrane (DRM) raft fractions. R24 treatment of the rat primary cerebellar cultures induced Lyn activation and tyrosine phosphorylation of Cbp. Treatment with anti‐ganglioside GD1b antibody also induced tyrosine phosphorylation. Furthermore, over‐expressions of Lyn and Cbp in Chinese hamster ovary (CHO) cells resulted in tyrosine 314 phosphorylation of Cbp, which indicates that Cbp is a substrate for Lyn. Immunoblotting analysis showed that the active form of Lyn and the Tyr314‐phosphorylated form of Cbp were highly accumulated in the DRM raft fraction prepared from the developing cerebellum compared with the DRM raft fraction of the adult one. In addition, Lyn and the Tyr314‐phosphorylated Cbp were highly concentrated in the growth cone fraction prepared from the developing cerebellum. Immunoelectron microscopy showed that Cbp and GAP‐43, a growth cone marker, are localized in the same vesicles of the growth cone fraction. These results suggest that Cbp functionally associates with gangliosides on growth cone rafts in developing cerebella. 相似文献
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Musashi Saigusa Mizuho Nishizawa Yutaka Shimizu 《Bioscience, biotechnology, and biochemistry》2013,77(9):1518-1527
Salmon myofibrillar protein (Mf) was investigated as a source of edible anti-inflammatory products. Peptides produced by stepwise digestion of Mf (without carbohydrate) with pepsin and trypsin had little effect on the secretion of inflammation-related compounds from lipopolysaccharide-stimulated RAW 264.7 macrophage cells. However, peptides prepared from Mf conjugated with alginate oligosaccharide (AO; 19 μg/mg protein) (dMSA) through the Maillard reaction in the presence of sorbitol significantly reduced the secretion of the pro-inflammatory mediators nitric oxide, tumor necrosis factor (TNF)-α and interleukin (IL)-6, as well as mRNA expression of TNF-α, IL-6, inducible nitric oxide synthase and cyclooxygenase-2. Additionally, dMSA inhibited acute inflammation in a carrageenan-induced model of paw edema in mice, but had no effect on natural killer cell cytotoxic activity or macrophage phagocytosis. These results suggest that fish Mf conjugated with AO may be a potential food material with anti-inflammatory function. 相似文献
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Hasegawa M Kawasaki A Yang K Fujimoto Y Masumoto J Breukink E Nuñez G Fukase K Inohara N 《The Journal of biological chemistry》2007,282(16):11757-11764
Nod1 is an intracellular protein that is involved in recognition of bacterial molecules and whose genetic variation has been linked to several inflammatory diseases. Previous studies suggested that the recognition core of Nod1 stimulatory molecules is gamma-D-glutamyl-meso-diaminopimelic acid (iE-DAP), but the identity of the major Nod1 stimulatory molecule produced by bacteria remains unknown. Here we show that bacteria produce lipophilic molecules capable of stimulating Nod1. Analysis of synthetic compounds revealed stereoselectivity of the DAP residue and that conjugation of lipophilic acyl residues specifically enhances the Nod1 stimulatory activity of the core iE-DAP. Furthermore, we demonstrate that lipophilic molecules induce and/or enhance the secretion of innate immune mediators from primary mouse mesothelial cells and human monocytic MonoMac6 cells, and this effect is mediated through Nod1. These results provide insight into the mechanism of immune recognition via Nod1, which might be useful in the design and testing of novel immunoregulators. 相似文献