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BACKGROUND: The D-alanyl-D-lactate (D-Ala-D-Lac) ligase is required for synthesis of altered peptidoglycan (PG) termini in the VanA phenotype of vancomycin-resistant enterococci (VRE), and the D-alanyl-D-serine (D-Ala-D-Ser) ligase is required for the VanC phenotype of VRE. Here we have compared these with the Escherichia coli D-Ala-D-Ala ligase DdlB for formation of the enzyme-bound D-alanyl phosphate, D-Ala(1)-PO(3)(2-) (D-Ala(1)-P), intermediate. RESULTS: The VanC2 ligase catalyzes a molecular isotope exchange (MIX) partial reaction, incorporating radioactivity from (14)C-D-Ser into D-Ala-(14)C-D-Ser at a rate of 0.7 min(-1), which approaches kinetic competence for the reversible D-Ala(1)-P formation from the back direction. A positional isotope exchange (PIX) study with the VanC2 and VanA ligases displayed a D-Ala(1)-dependent bridge to nonbridge exchange of the oxygen-18 label of [gamma-(18)O(4)]-ATP at rates of up to 0.6 min(-1); this exchange was completely suppressed by the addition of the second substrate D-Ser or D-Lac, respectively, as the D-Ala(1)-P intermediate was swept in the forward direction. As a third criterion for formation of bound D-Ala(1)-P, we conducted rapid quench studies to detect bursts of ADP formation in the first turnover of DdlB and VanA. With E. coli DdlB, there was a burst amplitude of ADP corresponding to 26-30% of the DdlB active sites, followed by the expected steady-state rate of 620-650 min(-1). For D-Ala-D-Lac and D-Ala-D-Ala synthesis by VanA, we measured a burst of 25-30% or 51% of active enzyme, respectively. CONCLUSIONS: These three approaches support the rapid (more than 1000 min(-1)), reversible formation of the enzyme intermediate D-Ala(1)-P by members of the D-Ala-D-X (where X is Ala, Ser or Lac) ligase superfamily. 相似文献
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The synthesis of capensifuranone (1) has been achieved by the application of developments for asymmetric conjugate addition reactions of organocopper reagents with nonracemic N-enoyl-4-phenyl-1,3-oxazolidinones for the preparation of 1,3-syn-dimethyl arrays. The assignment of relative and absolute stereochemistry of 1 has been made following the high-field NMR characterizations of synthetic diol derivatives. The previously unassigned C4 stereochemistry of 1 was determined to be of the (S)-configuration. The thermodynamic equilibration of capensifuranone and its C4 diastereomer has been examined. 相似文献
35.
The conjugate addition reactions of allylic stannanes have been investigated utilizing nonracemic N-enoyl-4-phenyl-1,3-oxazolidinones with Lewis acid precomplexation. 相似文献
36.
Conformationally locked isostere of phosphoSer-cis-Pro inhibits Pin1 23-fold better than phosphoSer-trans-Pro isostere 总被引:1,自引:0,他引:1
Wang XJ Xu B Mullins AB Neiler FK Etzkorn FA 《Journal of the American Chemical Society》2004,126(47):15533-15542
Stereoisomeric cis and trans substrate analogues for Pin1 were designed and synthesized. The central phosphoSer-Pro core of the Pin1 substrate was replaced by cis and trans amide isosteres in Ac-Phe-Phe-pSer-Psi[(Z and E)CH=C]-Pro-Arg-NH(2), 1 and 2, peptidomimetics. They were synthesized on solid phase in 17% yield for the cis analogue 1, and 16% yield for the trans analogue 2. A second trans amide isostere with a C-terminal N-methylamide 3 was synthesized in 7% yield. The protease-coupled Pin1 assay showed that all three compounds inhibited the Pin1 peptidyl-prolyl isomerase (PPIase) enzymatic activity. The cis isostere 1 was 23 times more potent (K(i) = 1.74 +/- 0.08 muM) than its trans counterpart 2 (K(i) = 40 +/- 2 muM) in competitive inhibition of Pin1. These results suggest that the catalytic site of Pin1 binds cis substrates more tightly in aqueous solution. Antiproliferative activity toward the A2780 human ovarian cancer cell line by the cis and trans analogues correlates with Pin1 inhibition results. 相似文献
37.
A procedure for determining the concentrations of dissolved chromium species in natural waters is described. Chromium(III) and chromium(VI), separated by co-precipitation with hydrated iron(III) oxide, and total dissolved chromium are determined separately by conversion to chromium(VI), extraction with APDC into MIBK and determination by a.a.s. The detection limit is 40 ng l?1 Cr. The dissolved chromium not amenable to separation and direct extraction is calculated by difference. In the waters investigated, total concentrations were relatively high (1–5 μg l?1) with Cr(VI) the predominant species in all areas sampled with one exception, where organically bound chromium was the major species. 相似文献
38.
Flibotte S Andrews HR Ball GC Beausang CW Beck FA Belier G Byrski T Curien D Dagnall PJ de France G Disdier D Duchêne G Finck C Haas B Hackman G Haslip DS Janzen VP Kharraja B Lisle JC Merdinger JC Mullins SM Nazarewicz W Radford DC Rauch V Savajols H Styczen J Theisen C Twin PJ Vivien JP Waddington JC Ward D Zuber K Åberg S 《Physical review letters》1993,71(26):4299-4302
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M. Piiparinen A. Ataç G. de Angelis S. Forbes N. Gjørup G. Hageman B. Herskind F. Ingebretsen H. Jensen D. Jerrestam H. Kusakari R. Lieder G. M. Marti S. Mullins J. Nyberg A. Santonocito H. Schnare G. Sletten K. Strahle M. Sugawara P. O. Tjøm A. Virtanen R. Wadsworth 《Zeitschrift für Physik A Hadrons and Nuclei》1992,343(3):367-368
The level scheme of 143 Eu has been extended to I=75/2 in an experiment with the NORDBALL Compton-suppressed Ge detector array and the 110 Pd(37 Cl, 4n) reaction. Most of the scheme shows irregular structure of multiparticle excitations. A strongly populated straight cascade of more than 10 stretched E2 transitions suggests the onset of collectivity. 相似文献