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- Cho, K. B., et al.
(författare)
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The substrate reaction mechanism of class III anaerobic ribonucleotide reductase
- 2001
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Ingår i: Journal of Physical Chemistry B. - : American Chemical Society (ACS). - 1089-5647 .- 1520-6106 .- 1520-5207. ; 105:27, s. 6445-6452
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Tidskriftsartikel (refereegranskat)abstract
- The substrate mechanism of class III anaerobic ribonucleotide reductase has been studied using quantum chemical methods. The study is based on the previously suggested mechanism for the aerobic class I enzyme, together with the recently determined X-ray structure of the anaerobic enzyme. The initial steps are similar in the mechanisms of these enzymes, but for the suggested rate-limiting steps there are key differences. In the class I enzyme, the 3 ' -keto group of the substrate is protonated in a step involving formation of a sulfur-sulfur bond between two cysteines, One of these cysteines is not present in the anaerobic enzyme. Instead, carbon dioxide is formed in this step from formate, which is present as a cofactor. In line with previous suggestions from experimental observations, the formate first forms a formyl radical. The next step, where the formyl radical protonates the 3 ' -keto group of the substrate, is suggested to be rate limiting with a calculated total barrier of 19.9 kcal/mol, in reasonable agreement with the experimental rate-limiting barrier of 17 kcal/mol. Zero-point and entropy effects are found to be quite significant in lowering the barrier. The mechanism for the entire cycle is discussed in relation to known experimental facts.
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- Engstrom, Maria, et al.
(författare)
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Hydrogen bonding to tyrosyl radical analyzed by ab initio g-tensor calculations
- 2000
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Ingår i: Journal of Physical Chemistry A. - : American Chemical Society (ACS). - 1089-5639 .- 1520-5215. ; 104:21, s. 5149-5153
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Tidskriftsartikel (refereegranskat)abstract
- Hydrogen bonding to the tyrosyl radical in ribonucleotide reductase (RNR) has been simulated by a complex between the phenoxyl radical and a water molecule. Multiconfigurational self-consistent field linear response theory was used to calculate the g-tensor of the isolated phenoxyl radical and of the phenoxyl-water model. The relevance of the model was motivated by the fact that spin density distributions and electron paramagnetic resonance (EPR) spectra of the phenoxyl and tyrosyl radicals are very similar. The calculated g-tensor anisotropy of the phenoxyl radical was comparable with experimental findings for tyrosyl in those RNRs where the H-bond is absent: g(x) = 2.0087(2.0087), g(y) = 2.0050(2.0042), and g(z) = 2.0025(2.0020), where the tyrosyl radical EPR data from Escherichia coli RNR are given in parentheses. The hydrogen bonding models reproduced a shift toward a lower g(x) value that was observed experimentally for mouse and herpes simplex virus RNR where the H-bond was detected by electron-nuclear double resonance after deuterium exchange. This decrease could be traced to lower angular momentum and spin-orbit coupling matrix elements between the ground B-2(1) and the first excited B-2(2) states (oxygen lone-pair n to pi(SOMO) excitation) upon hydrogen bonding in a linear configuration. The g(x) value was further decreased by hydrogen bonding in bent configurations due to a blue shift of this excitation.
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- Lindskog, M, et al.
(författare)
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Monitoring intracellular metabolites in neuroblastoma with 1H NMR spectroscopy: effects of growth factor withdrawal and modulation of lipid metabolism
- 2004
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Ingår i: SPECTROSCOPY-AN INTERNATIONAL JOURNAL. - : Hindawi Limited. - 0712-4813. ; 18:2, s. 123-132
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- 1H NMR spectroscopy has previously been employed to detect and monitor changes in the lipid metabolism of neuroblastoma cells upon cytotoxic treatment. Here, we addressed the question whether altered growth conditions, by presence or absence of serum, would impact on the metabolites detectable with1H NMR spectroscopy. Chronic serum deprivation of SH‒SY5Y human neuroblastoma cells resulted in a decrease in the intracellular content of several metabolites, in particular total choline. This metabolic effect was paralleled by significant growth inhibition. In addition, we investigated the potential functional origin of intracellular1H NMR visible lipids in SH‒SY5Y cells. A drop in lipid methylene protons could be observed shortly after serum‒withdrawal. Contrary, removal of lipoproteins from the serum led to a pronounced increase in intracellular lipids, as did inhibition of de novo sterol synthesis by lovastatin. In conclusion, we demonstrate that intracellular total choline in neuroblastoma cells in vitro is highly dependent on the availability of growth factors. Furthermore, we show that1H NMR visible lipids decrease upon serum‒withdrawal but are accumulated when cholesterol supply is abrogated. The biological and potential clinical implications of these findings are discussed.
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