Simultaneous acquisition of 2D and 3D solid-state NMR experiments for sequential assignment of oriented membrane protein samples

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Simultaneous acquisition of 2D and 3D solid-state NMR experiments for sequential assignment of oriented membrane protein samples

Abstract

We present a new method called DAISY (Dual Acquisition orIented ssNMR spectroScopY) for the simultaneous acquisition of 2D and 3D oriented solid-state NMR experiments for membrane proteins reconstituted in mechanically or magnetically aligned lipid bilayers. DAISY utilizes dual acquisition of sine and cosine dipolar or chemical shift coherences and long living 15N longitudinal polarization to obtain two multi-dimensional spectra, simultaneously. In these new experiments, the first acquisition gives the polarization inversion spin exchange at the magic angle (PISEMA) or heteronuclear correlation (HETCOR) spectra, the second acquisition gives PISEMA-mixing or HETCOR-mixing spectra, where the mixing element enables inter-residue correlations through 15Nâ??15N homonuclear polarization transfer. The analysis of the two 2D spectra (first and second acquisitions) enables one to distinguish 15Nâ??15N inter-residue correlations for sequential assignment of membrane proteins. DAISY can be implemented in 3D experiments that include the polarization inversion spin exchange at magic angle via I spin coherence (PISEMAI) sequence, as we show for the simultaneous acquisition of 3D PISEMAIâ??HETCOR and 3D PISEMAIâ??HETCOR-mixing experiments.



Source: Journal of Biomolecular NMR