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81.
Function of calmodulin in postsynaptic densities. II. Presence of a calmodulin- activatable protein kinase activity 下载免费PDF全文
Because the calmodulin in postsynaptic densities (PSDs) activates a cyclic nucleotide phosphodiesterase, we decided to explore the possibility that the PSD also contains a calmodulin-activatable protein kinase activity. As seen by autoradiographic analysis of coomassie blue-stained SDS polyacrylamide gels, many proteins in a native PSD preparation were phosphorylated in the presence of [γ-(32)P]ATP and Mg(2+) alone. Addition of Ca(2+) alone to the native PSD preparation had little or no effect on phosphorylation. However, upon addition of exogenous calmodulin there was a general increase in background phosphorylation with a statistically significant increase in the phosphorylation of two protein regions: 51,000 and 62,000 M(r). Similar results were also obtained in sonicated or freeze thawed native PSD preparations by addition of Ca(2+) alone without exogenous calmodulin, indicating that the calmodulin in the PSD can activate the kinase present under certain conditions. The calmodulin dependency of the reaction was further strengthened by the observed inhibition of the calmodulin-activatable phosphorylation, but not of the Mg(2+)-dependent activity, by the Ca(2+) chelator, EGTA, which also removes the calmodulin from the structure (26), and by the binding to calmodulin of the antipsychotic drug chlorpromazine in the presence of Ca(2+). In addition, when a calmodulin-deficient PSD preparation was prepared (26), sonicated, and incubated with [γ-(32)P]ATP, Mg(2+) and Ca(2+), one could not induce a Ca(2+)-stimulation of protein kinase activity unless exogenous calmodulin was added back to the system, indicating a reconstitution of calmodulin into the PSD. We have also attempted to identify the two major phosphorylated proteins. Based on SDS polyacrylamide gel electrophoresis, it appears that the major 51,000 M(r) PSD protein is the one that is phosphorylated and not the 51,000 M(r) component of brain intermediate filaments, which is a known PSD contaminant. In addition, papain digestion of the 51,000 M(r) protein revealed multiple phosphorylation sites different from those phosphorylated by the Mg(2+)-dependent kinase(s). Finally, although the calmodulin-activatable protein kinase may phosphorylate proteins I(a) and I(b), the cyclic AMP-dependent protein kinase, which definitely does phosphorylate protein I(a) and I(b) and is present in the PSD, does not phosphorylate the 51,000 and 62,000 M(r) proteins, because specific inhibition of this kinase has no effect on the levels of the phosphorylation of these latter two proteins. 相似文献
82.
Abortive initiation and long ribonucleic acid synthesis 总被引:11,自引:0,他引:11
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G. Sachs K. Munson V. N. Balaji D. Aures-Fischer S. J. Hersey K. Hall 《Journal of bioenergetics and biomembranes》1989,21(5):573-588
The gastric H+ + K+ ATPase is a member of the phosphorylating class of transport ATPase. Based on sequence homologies and CHO content, there may be ab subunit associated with the catalytic subunit of the H+ + K+ ATPase. Its function, if present, is unknown. The pump catalyzes a stoichiometric exchange of H+ for K+, but is also able to transport Na+ in the forward direction. This suggests that the transport step involves hydronium rather than protons. The initial binding site is likely to contain a histidine residue to account for the high affinity of the cellular site. The extracellular site probably lacks this histidine, so that a low affinity for hydronium allows release into a solution of pH 0.8. Labelling with positively charge, luminally reactive reagents that block ATPase and pump activity has shown that a region containing H5 and H6 and the intervening luminal loop is involved in necessary conformational changes for normal pump activity. The calculated structure of this loop shows the presence of ana helical,b turn, andb strand sector, with negative charges close to the membrane domain. This sector provides a possible site of interaction of drugs with the H+ + K+ ATPase, and may be part of the K+ pathway in the enzyme.Emory University, Atlanta, Georgia. 相似文献
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Linda Munson 《Zoo biology》1993,12(1):105-124
Knowledge of the diseases of cheetahs is essential to prevent and treat conditions that can modulate fertility and longevity. Toward this aim, a comprehensive pathology survey was conducted under a directive from the Cheetah Species Survival Plan. To date, 31 adult cheetahs and nine cubs from 16 zoological parks have been evaluated. Also, liver biopsies from 67 female cheetahs from 22 zoological parks were examined. Veno-occlusive disease (VOD) affected 82% of deceased cheetahs and 51% of live female cheetahs, and was the cause of death in nine cheetahs. Glomerulosclerosis and nephrosclerosis affected 84% and 39% of the population, respectively, and caused renal failure in eight cheetahs. The severity of VOD and glomerulosclerosis increased with age, and was not associated with infertility. Chronic gastritis was noted in 91% of the study population, and 95% of these cases also had spiral bacteria. Feline infectious peritonitis caused the death of two cheetahs. Male cheetahs had testicular degeneration, atrophy, and/or spermatogenic arrest, but these cheetahs also had severe systemic illness. Most females did not have reproductive tract lesions that would cause infertility, including those with parovarian cysts. Ovarian histology suggested that infertile cheetahs were not ovulating. Most cubs died from pneumonia or other systemic infections. The results of this study indicate that serious diseases are prevalent in the North American cheetahs, but these diseases do not appear to be the cause of infertility in the population. However, these diseases do limit the life span and well-being of cheetahs in captivity. Further research is needed to elucidate the causes of these diseases. © 1993 Wiley-Liss, Inc. 相似文献
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A report of the 23rd Protein Symposium 'Proteins in Motion', Boston, USA, 23-27 July 2009. 相似文献
89.
The cell envelope fraction of Salmonella typhimurium contains an enzyme system which catalyzes transfer of 3-deoxyoctulosonate (KDO) from CMP-KDO to an incomplete, KDO-deficient precursor of lipid A. The enzyme system is firmly membrane-bound, but has been solubilized by treatment with nonionic detergent at alkaline pH and partially purified. Both the particulate and partially purified fractions catalyzed formation of a single reaction product containing 2 residues of KDO. Periodate oxidation of the purified product permitted tentative identification of the KDO disaccharide structure as KDO2-4KDO. 相似文献
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