Two-field transverse relaxation-optimized spectroscopy for the study of large biomolecules – An in silico investigation

Nicolas Bolik-Coulon,Philippe Pelupessy,Guillaume Bouvignies,Fabien Ferrage

doi:10.1016/j.jmro.2020.100007

Abstract

Biomolecular NMR spectroscopy has greatly benefited from the development of TROSY-type pulse sequences, in pair with specific labeling. The selection of spin operators with favorable relaxation properties has led to an increase in the resolution and sensitivity of spectra of large biomolecules. However, nuclei with a large chemical shift anisotropy (CSA) contribution to relaxation can still suffer from large linewidths at conventional magnetic fields (higher than 9 T). Here, we introduce the concept of two-field TROSY (2F-TROSY) where the chemical shifts of nuclei with large CSA is labeled at low fields (ca. 2 T) dramatically reducing the contribution of CSA to relaxation. Signal detection is performed at high field (> 9 T) on a nucleus with efficient TROSY interference to yield high resolution and sensitivity. We use comprehensive numerical simulations to demonstrate the power of this approach on aromatic 13C-19F spin pairs for which a TROSY pulse sequence has recently been published. We predict that the 2F-TROSY experiment shall yield good quality spectra for large proteins (global tumbling correlation times as high as 100 ns) with one order of magnitude higher sensitivity than the single-field experiment.

Highlights

Nuclear Magnetic Resonance (NMR) is a powerful tool to investigate the structure, dynamics and function of complex biomolecular systems at atomic resolution
The methyl-transverse relaxation-optimized spectroscopy (TROSY) experiment is based on the cancellation between intra-methyl dipole-dipole (DD) interactions while the NH-TROSY exploits interference between DD and Chemical Shift Anisotropy (CSA) interactions.[8,11,12,13]
We have introduced the concept of two-field transverse relaxation-optimized spectroscopy (2FTROSY)

Summary

INTRODUCTION

Nuclear Magnetic Resonance (NMR) is a powerful tool to investigate the structure, dynamics and function of complex biomolecular systems at atomic resolution. The set of two-spin TROSY pulse sequences used for the study of biomolecules has mainly been applied on pairs of the type X-1H (X=backbone-15N in proteins or aromatic-15N and 13C in proteins and nucleic acids).[4,5,6,7] In these spin systems, the CSA of the protons is either small or comparable to the amplitude of the DD interaction at magnetic fields currently accessible (between 9 and 28 T) and leads to field-dependence of the proton transverse relaxation rate usually less pronounced than for the relaxation of backbone-15N and aromatic-13C nuclei.[15] the optimal field for the associated TROSY experiment depends mostly on the relaxation properties of the heteronucleus This is not the case in the recently developed two-dimensional 13C-19F-TROSY experiment for the study of 19F-labeled protein aromatic sidechains and nucleic acid bases.16,1718. We expect that 2F-TROSY will enhance NMR of large biomolecules by making a diversity of new spin systems accessible, starting with 13C-19F pairs in aromatic side-chains

THEORY AND CALCULATIONS

RESULTS AND DISCUSSION

CONCLUSION