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11.
Herje Schagerl?f Carina Nilsson Lo Gorton Folke Tjerneld Henrik St?lbrand Arieh Cohen 《Analytical and bioanalytical chemistry》2009,394(7):1977-1984
We present a novel method for investigating subsite-substrate interactions of glycoside hydrolases and the determination of
the oligosaccharide cleavage point based on the analysis of the hydrolysis products produced in the presence of 18O-labelled water. Conventional techniques for such determination of the hydrolysis pattern call for the chemical modification
of the substrate, whereas the method presented makes it possible to use natural substrates, utilising the selectivity and
sensitivity of mass spectrometry. This method is very useful for the detection and analysis of enzyme-catalysed hydrolysis,
provided that the conditions are chosen where 18O incorporation without the presence of the enzyme is absent or undetectable. Such conditions were found and used in incubations
of cellopentaose with the well-characterised endoglucanase Cel5A from Bacillus agaradhaerens. We were able to confirm that the preferred glycoside bond to be hydrolysed is the third one counting from the non-reducing
end of the cellopentaose. Thus, cellopentaose prefers to bind from the –3 to the +2 subsites, which is in accordance with
published crystallographic data. The main advantage of the method presented is that there is no need for a priori chemical
modification/labelling of oligosaccharide substrates, which are processes that can disturb the enzyme–substrate interaction.
From 18O incorporation we could demonstrate that the enzyme also has an oxygen-exchange activity on cellotriose and cellobiose. This
is in agreement with the mechanism for transglycosylation and indicates that it is possible for the enzyme to perform such
reactions. 相似文献
12.
Cohen A Schagerlöf H Nilsson C Melander C Tjerneld F Gorton L 《Journal of chromatography. A》2004,1029(1-2):87-95
A series of celloendoglucanases: Bacillus agaradhaerens Cel 5a, Humicola insolens Cel 5a, H. insolens Cel 7b, H. insolens Cel 45a, Trichoderma reesei Cel 7b, and T. reesei Cel 45a were used to hydrolyse carboxymethylcellulose (CMC) and the hydrolysis products were investigated with a novel liquid chromatography-mass spectrometry (LC-MS) method. Separation was achieved using a graphitised carbon chromatographic column which allowed the use of electrospay compatible eluents. Analysis of the compounds produced during enzyme hydrolysis of CMC is used to understand enzyme selectivities and substitution pattern of CMC. Conventional high-performance anion-exchange chromatography (HPAEC)-pulsed amperometric detection (PAD), size-exclusion chromatography (SEC)-refractive index (RI) detection, and reducing end analysis are also used to analyse enzyme-hydrolysed CMC. The LC-MS method presented allows for a more detailed investigation of hydrolysis products, which facilitates characterisation of both enzymes and substrates. 相似文献
13.
Ragnhild B. Strand Trygve Helgerud Tina Solvang Amund Dolva Christian A. Sperger Anne Fiksdahl 《Tetrahedron: Asymmetry》2012,23(18-19):1350-1359
Screening studies of new chiral imidazolium and triazolium based NHC salts I–VIII as ligands in asymmetric organometallic catalysis and as organocatalysts showed that these catalysts efficiently promoted the reactions. Moderate enantioselectivities (55–57% ee) were obtained in the asymmetric Cu-NHC catalysed conjugate additions of diethylzinc to cyclohexenone, in accordance with most previous studies. The chiral induction afforded in the gold(I)-NHC catalysed cyclopropanation reactions was low (<28% ee). However, these results represent the first reported chiral gold(I)-NHC catalysed olefin cyclopropanation. The NHC-organocatalysed asymmetric cross-annulation of cinnamaldehyde and trifluoroacetophenone gave lower enantioselectivity (<50% ee) but higher yields of the γ-lactone product relative to previous reports. The enantioselectivities obtained varied considerably, even within a group of structurally closely related NHCs. This study demonstrates the challenge of designing NHCs with a general ability to induce asymmetry in a broader range of reactions. 相似文献
14.
Park Y Whitaker RD Nap RJ Paulsen JL Mathiyazhagan V Doerrer LH Song YQ Hürlimann MD Szleifer I Wong JY 《Langmuir : the ACS journal of surfaces and colloids》2012,28(15):6246-6255
The detection of superparamagnetic nanoparticles using NMR logging has the potential to provide enhanced contrast in oil reservoir rock formations. The stability of the nanoparticles is critical because the NMR relaxivity (R(2) ≡ 1/T(2)) is dependent on the particle size. Here we use a molecular theory to predict and validate experimentally the stability of citric acid-coated/PEGylated iron oxide nanoparticles under different pH conditions (pH 5, 7, 9, 11). The predicted value for the critical surface coverage required to produce a steric barrier of 5k(B)T for PEGylated nanoparticles (MW 2000) was 0.078 nm(-2), which is less than the experimental value of 0.143 nm(-2), implying that the nanoparticles should be stable at all pH values. Dynamic light scattering (DLS) measurements showed that the effective diameter did not increase at pH 7 or 9 after 30 days but increased at pH 11. The shifts in NMR relaxivity (from R(2) data) at 2 MHz agreed well with the changes in hydrodynamic diameter obtained from DLS data, indicating that the aggregation behavior of the nanoparticles can be easily and quantitatively detected by NMR. The unexpected aggregation at pH 11 is due to the desorption of the surface coating (citric acid or PEG) from the nanoparticle surface not accounted for in the theory. This study shows that the stability of the nanoparticles can be predicted by the theory and detected by NMR quantitatively, which suggests the nanoparticles to be a possible oil-field nanosensor. 相似文献
15.
The molecular structures of gaseous tetrafluoro-p-benzoquinone (p-fluoranil) and tetramethyl-p-benzoquinone (duroquinone) have been investigated by electron diffraction. Except for the methyl group hydrogen atoms, the molecules are planar to within experimental error, but small deviations from planarity are completely compatible with the data. Values for the geometrical parameters (radistances and rα with parenthesized uncertainties of 2σ including estimated uncertainty in the electron wavelength and correlation effects, are as follows. Tetrafluoro-p-benzoquinone: D2h symmetry (assumed); r(C0) = 1.211(6) Å, r(CC) = 1.339(12) Å, r(C-C) = 1.489(5') Å, r(C-F) = 1.323(5) Å, ∠C-C-C = 116.8(7)° and ∠C-C-F = 116.1(7)°. Tetramethyl-p-benzoquinone: C2h symmetry (assumed);r(C-H) = 1.102(18) Å, r(CO) = 1.229(8) Å, r(CC) = 1.352(8) Å, r(Csp2-Csp2) = 1.491(11) Å, r(Csp2-Csp3) = 1.504(12) A, ∠C-CO-C = 120.8(8)°. ∠C-C-CH3 = 116.1(8)°, ∠C-C-H = 110.5(34)° and α1 = α2 (methyl torsion = 30° (assumed). 相似文献