# Orientation Selection in Proton-Detected Magic-Angle Spinning Torsion Angle Experiments

## TL;DR

This paper improves the accuracy of determining protein backbone angles using NMR by addressing issues caused by cross-polarization in experiments.

## Contribution

The study introduces optimized cross-polarization conditions to enhance torsion angle accuracy in MAS NMR experiments.

## Key findings

- Optimized CP conditions improve agreement between NMR torsion angles and X-ray data for chicken α-spectrin SH3.
- An unexpected backbone angle was found for influenza A M2 I32 residue, aligning with TALOS-N predictions.
- The proposed method reduces orientation bias in torsion angle determination using pseudo-4D experiments.

## Abstract

Determination of torsion angles via recoupling of backbone
HC and
HN dipolar interactions is a well-known method in magic-angle spinning
NMR spectroscopy. Torsion angle values can be obtained by comparing
simulated and experimental signals, either in the frequency or time
domains. Typically, all molecular orientations are assumed to have
identical detected amplitudes at zero recoupling time. The changes
in these amplitudes during the recoupling period define the dipolar
coupling values and the torsion angles. Experimentally, however, orientations
may exhibit different detected amplitudes due to additional cross-polarization
(CP) blocks that connect different spins in multidimensional experiments.
We numerically and experimentally investigate how CP blocks bias backbone
φ torsion angle determination and propose CP conditions that
minimize this effect, thereby improving accuracy. Applying these conditions
in pseudo-4D (H)­CANH experiments yields improved agreement of the
extracted angles with X-ray crystallographic data for microcrystalline
chicken α-spectrin SH3. For the influenza A M2 membrane protein,
we identify an unexpected backbone dihedral angle for the I32 residue,
which is consistent with TALOS-N predictions but deviates from ideal
α-helical transmembrane geometry.