Improved method for unambiguous amino acid side-chain 1H and 13C resonance assignment |
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Authors: | Löhr Frank Betz Marco Rüterjans Heinz |
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Institution: | Institut für Biophysikalische Chemie, Zentrum für Biomolekulare Magnetische Resonanz, Johann Wolfgang Goethe-Universit?t, Marie Curie-Strasse 9, D-60439 Frankfurt am Main, Germany. murph@bpc.uni-frankfurt.de |
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Abstract: | Side-chain proton and carbon-13 resonance assignments of 13C;15N]-enriched proteins usually rely on combinations of several multi-dimensional experiments. Here, we describe a four-dimensional pulse sequence, H(C)C-COSY-TOCSY-(CACO)NH, which provides the information required to assign completely aliphatic side-chain resonance frequencies. As in widely used HCC(CO)NH-TOCSY experiments, problems due to spectral crowding are alleviated by exploiting the dispersion of backbone amide 1H and 15N signals. The modification introduced here allows signals from different side-chains to be distinguished even in the case of overlap in the 1H(N)-15N plane of the spectra. For illustration, the new method is applied to two proteins with molecular masses of 11 and 23 kDa. |
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Keywords: | NMR 1H NMR 13C NMR 15N NMR CC‐TOCSY CC‐COSY triple resonance spectroscopy RNase T1 xylanase |
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