J-based 2D homonuclear and heteronuclear correlation in solid-state proteins

Abstract

Scalar-based two-dimensional heteronuclear experiments are reported for NCO and NCA chemical shift correlation in the solid state. In conjunction with homonuclear CACO correlation, these experiments form a useful set for tracing connectivities and assigning backbone resonances in solid-state proteins. The applicability of this approach is demonstrated on two proteins, the β 1 immunoglobulin binding domain of protein G at 9.4 T and reassembled thioredoxin at 14.1 T, using different decoupling conditions and MAS frequencies. These constant-time J-based correlation experiments exhibit increased resolution in the indirect dimension owing to homonuclear and heteronuclear decoupling, and because the indirect evolution and transfer periods are combined into a single constant time interval, this increased resolution is not obtained at the cost of sensitivity. These experiments are also shown to be compatible with in-phase anti-phase (IPAP) selection, giving increased resolution in the directly detected dimension.

High resolution 13C-detected solid-state NMR spectroscopy of a deuterated protein

“…Our results show significant improvement in T 2 relaxation times even with low proton-decoupling power that will make it possible to perform J -based SSNMR experiments (Chen et al 2006, 2007) to improve resolution of non-crystalline samples. The improvement in the T 2 relaxation times and resolution will facilitate the sequential backbone assignments of large systems in which peak separation represents a big challenge, and exchangeable protons or water protons will enhance the selective detection of polar residues or residues that are close to water molecules.…”

Reduction of spin diffusion artifacts from 2D zfr-INADEQUATE MAS NMR spectra

“…Another alternative that has been explored by Mueller and others [38,39] involves using heteronuclear decoupling to help remove the artifacts that arise from the residual dipolar interactions. We have not compared our experiments to these heteronuclear versions and this may well be worth future study.…”

Chapter 2. Solid‐State NMR in Drug Discovery and Development

“…85,86 A range of 3D heteronuclear J-based sequences designed for proteins have been developed to mimic experiments used in solution-state protein NMR; these include constant-time NCA, NCO, NCACO, NCOCA and CANCO experiments for direct tracing of the backbone of 13 C and 15 N-labeled proteins. 87,88 The measurement of the CST for a nucleus provides another important parameter that can be related to structure. While CSTs can be measured by direct observation of static lineshapes or MAS spinning sideband manifolds, these approaches are limited to relatively simple spectra containing only a few resonances.…”