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排序方式: 共有76条查询结果,搜索用时 93 毫秒
1.
Purification and characterization of glucose dehydrogenase from the thermoacidophilic archaebacterium Thermoplasma acidophilum. 下载免费PDF全文
Glucose dehydrogenase was purified to homogeneity from the thermoacidophilic archaebacterium Thermoplasma acidophilum. The enzyme is a tetramer of polypeptide chain Mr 38,000 +/- 3000, it is catalytically active with both NAD+ and NADP+ cofactors, and it is thermostable and remarkably resistant to a variety of organic solvents. The amino acid composition was determined and compared with those of the glucose dehydrogenases from the archaebacterium Sulfolobus solfataricus and the eubacteria Bacillus subtilis and Bacillus megaterium. The N-terminal amino acid sequence of the Thermoplasma acidophilum enzyme was determined to be: (S/T)-E-Q-K-A-I-V-T-D-A-P-K-G-G-V-K-Y-T-T-I-D-M-P-E. 相似文献
2.
Models of proteolysis of oligomeric enzymes and their applications to the trypsinolysis of citrate synthases. 下载免费PDF全文
A simple statistical approach was used to generate predictive models of the proteolysis of multisubunit enzymes in order to correlate the loss of enzyme activity with the loss of native subunit. The models were applied to the trypsinolysis of the citrate synthases of pig heart, Bacillus megaterium and Escherichia coli. With the dimeric citrate synthases (pig heart and B. megaterium) trypsinolysis of one of the subunits appears to destroy the activity of the whole enzymic molecule. The hexameric E. coli citrate synthase behaves like a trimer of dimeric units, each of the dimers behaving similarly to the B. megaterium and pig heart enzymes. Palmitoyl-CoA is required for the trypsinolysis of pig heart citrate synthase, and at relatively high concentrations of this compound trypsinolysis of one subunit leaves the other subunit fully active. Palmitoyl-CoA is not required for the trypsinolysis of the other citrate synthases, and high concentrations of this metabolite do not affect the correlation of proteolysis with inactivation of these enzymes. 相似文献
3.
The discovery of an additional step in the progression of an enzyme from the active to inactive state under the influence of temperature has led to a better match with experimental data for all enzymes that follow Michaelis–Menten kinetics, and to an increased understanding of the process. The new model of the process, the Equilibrium Model, describes an additional mechanism by which temperature affects the activity of enzymes, with implications for ecological, metabolic, structural, and applied studies of enzymes. 相似文献
4.
Heath C Posner MG Aass HC Upadhyay A Scott DJ Hough DW Danson MJ 《The FEBS journal》2007,274(20):5406-5415
The aerobic archaea possess four closely spaced, adjacent genes that encode proteins showing significant sequence identities with the bacterial and eukaryal components comprising the 2-oxoacid dehydrogenase multi-enzyme complexes. However, catalytic activities of such complexes have never been detected in the archaea, although 2-oxoacid ferredoxin oxidoreductases that catalyze the equivalent metabolic reactions are present. In the current paper, we clone and express the four genes from the thermophilic archaeon, Thermoplasma acidophilum, and demonstrate that the recombinant enzymes are active and assemble into a large (M(r) = 5 x 10(6)) multi-enzyme complex. The post-translational incorporation of lipoic acid into the transacylase component of the complex is demonstrated, as is the assembly of this enzyme into a 24-mer core to which the other components bind to give the functional multi-enzyme system. This assembled complex is shown to catalyze the oxidative decarboxylation of branched-chain 2-oxoacids and pyruvate to their corresponding acyl-CoA derivatives. Our data constitute the first proof that the archaea possess a functional 2-oxoacid dehydrogenase complex. 相似文献
5.
Lamble HJ Heyer NI Bull SD Hough DW Danson MJ 《The Journal of biological chemistry》2003,278(36):34066-34072
The hyperthermophilic Archaeon Sulfolobus solfataricus metabolizes glucose by a non-phosphorylative variant of the Entner-Doudoroff pathway. In this pathway glucose dehydrogenase and gluconate dehydratase catalyze the oxidation of glucose to gluconate and the subsequent dehydration of gluconate to 2-keto-3-deoxygluconate. 2-Keto-3-deoxygluconate (KDG) aldolase then catalyzes the cleavage of 2-keto-3-deoxygluconate to glyceraldehyde and pyruvate. The gene encoding glucose dehydrogenase has been cloned and expressed in Escherichia coli to give a fully active enzyme, with properties indistinguishable from the enzyme purified from S. solfataricus cells. Kinetic analysis revealed the enzyme to have a high catalytic efficiency for both glucose and galactose. KDG aldolase from S. solfataricus has previously been cloned and expressed in E. coli. In the current work its stereoselectivity was investigated by aldol condensation reactions between D-glyceraldehyde and pyruvate; this revealed the enzyme to have an unexpected lack of facial selectivity, yielding approximately equal quantities of 2-keto-3-deoxygluconate and 2-keto-3-deoxygalactonate. The KDG aldolase-catalyzed cleavage reaction was also investigated, and a comparable catalytic efficiency was observed with both compounds. Our evidence suggests that the same enzymes are responsible for the catabolism of both glucose and galactose in this Archaeon. The physiological and evolutionary implications of this observation are discussed in terms of catalytic and metabolic promiscuity. 相似文献
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Moore V Kanu A Byron O Campbell G Danson MJ Hough DW Crennell SJ 《Extremophiles : life under extreme conditions》2011,15(3):327-336
Using citrate synthase from the hyperthermophile Pyrococcus furiosus (PfCS) as our test molecule, we show through guanidine hydrochloride-induced unfolding that the dimer separates into folded,
but inactive, monomers before individual subunit unfolding takes place. Given that forces across the dimer interface are vital
for thermostability, a robust computational method was derived that uses the University of Houston Brownian Dynamics (UHBD)
program to calculate both the hydrophobic and electrostatic contribution to the dimerisation energy at 100°C. The results
from computational and experimental determination of the lowered stability of interface mutants were correlated, being both
of the same order of magnitude and placing the mutant proteins in the same order of stability. This computational method,
optimised for hyperthermophilic molecules and tested in the laboratory, after further testing on other examples, could be
of widespread use in the prediction of thermostabilising mutations in other oligomeric proteins for which dissociation is
the first step in unfolding. 相似文献
9.
Those aerobic archaea whose genomes have been sequenced possess a single 4-gene operon that, by sequence comparisons with Bacteria and Eukarya, appears to encode the three component enzymes of a 2-oxoacid dehydrogenase multienzyme complex. However, no catalytic activity of any such complex has ever been detected in the Archaea. In the current paper, we have cloned and expressed the first two genes of this operon from the thermophilic archaeon, Thermoplasma acidophilum. We demonstrate that the protein products form an alpha2beta2 hetero-tetramer possessing the decarboxylase catalytic activity characteristic of the first component enzyme of a branched-chain 2-oxoacid dehydrogenase multienzyme complex. This represents the first report of the catalytic function of these putative archaeal multienzyme complexes. 相似文献
10.
An investigation has been carried out into gluconate dehydratase from the hyperthermophilic Archaeon Sulfolobus solfataricus. The enzyme has been purified from cell extracts of the organism and found to be responsible for both gluconate and galactonate dehydratase activities. It was shown to be a 45 kDa monomer with a half-life of 41 min at 95 degrees C and it exhibited similar catalytic efficiency with both substrates. Taken alongside the recent work on glucose dehydrogenase and 2-keto-3-deoxygluconate aldolase, this report clearly demonstrates that the entire non-phosphorylative Entner-Doudoroff pathway of S. solfataricus is promiscuous for the metabolism of both glucose and galactose. 相似文献