High-Resolution Four-Dimensional ~1H-~(13)C NOE Spectroscopy using Methyl-TROSY,Sparse Data Acquisition,and Multidimensional Decomposition

摘要

An approach for recording four-dimensional (4D) methyl ~1H-~(13)C-~(13)C-~1H NOESY spectra with high resolution and sensitivity is presented and applied to Malate Synthase G (723 residues,82 kDa).Sensitivity and resolution have been optimized using a highly deuterated,methyl-protonated sample in concert with methyl-TROSY,sparse data sampling in the three indirect dimensions,and 4D spectral reconstruction using multidimensional decomposition (MDD).A sparse data acquisition protocol is introduced that ensures that sufficiently long indirect acquisition times can be employed to exploit the decreased relaxation rates associated with methyl-TROSY,without increasing the duration of the 4D experiment beyond acceptable measurement times.In this manner,only a fraction (approx30%) of the experimental data that would normally be needed to achieve a spectrum of high resolution is acquired.The reconstructed 4D spectrum is of similar resolution and sensitivity to three-dimensional (3D) ~(13)C-edited NOE spectra,is straightforward to analyze,and resolves ambiguities that emerge when 3D data sets only are considered.