UTOPIA NMR: activating unexploited magnetization using interleaved low-gamma detection

Abstract

A growing number of nuclear magnetic resonance (NMR) spectroscopic studies are impaired by the limited information content provided by the standard set of experiments conventionally recorded. This is particularly true for studies of challenging biological systems including large, unstructured, membrane-embedded and/or paramagnetic proteins. Here we introduce the concept of unified time-optimized interleaved acquisition NMR (UTOPIA-NMR) for the unified acquisition of standard high-γ (e.g. 1H) and low-γ (e.g. 13C) detected experiments using a single receiver. Our aim is to activate the high level of polarization and information content distributed on low-γ nuclei without disturbing conventional magnetization transfer pathways. We show that using UTOPIA-NMR we are able to recover nearly all of the normally non-used magnetization without disturbing the standard experiments. In other words, additional spectra, that can significantly increase the NMR insights, are obtained for free. While we anticipate a broad range of possible applications we demonstrate for the soluble protein Bcl-xL (ca. 21 kDa) and for OmpX in nanodiscs (ca. 160 kDa) that UTOPIA-NMR is particularly useful for challenging protein systems including perdeuterated (membrane) proteins.

The power, pitfalls and potential of the nanodisc system for NMR-based studies

“…Alternatively, NMRoptimized, smaller MSP constructs (see Section 'Choosing the MSP construct') can be used to sufficiently increase molecular tumbling and allow the usage of regular 13 C-13 C transfer sequences . It has also been successfully shown that TOCSY-based 13 C-13 C transfer, combined with 13 C detection, can be used to study the transmembrane region of OmpX in MSDP1D1 NDs (Viegas et al, 2016). Remarkably, when using unified time-optimized interleaved acquisition (UTOPIA) setups this information can be obtained for free during the acquisition of a 3D NOESY-TROSY experiment (Viegas et al, 2016).…”

“…One of the greatest potentials of using NMR spectroscopy to study MPs incorporated into nanodiscs is the possibility of determining their 3D structure in solution, in a closeto-native and active conformation. Although the molecular weight of the MP-ND complex is high, it should still be under 200 kDa and thus suitable for being studied by modern NMR techniques (Shenkarev et al, 2010a;Klammt et al, 2012;Opella 2013;Viegas et al, 2016). The first MP structure determined by NMR and using the ND technology was of the 180-residues bacterial outer-membrane protein OmpX (PDB code: 2M06) .…”

Parallel NMR spectroscopy with simultaneous detection of 1H and 19F nuclei

“…Examples of the three types of experiments involving two receivers: (a) interleaved experiments, e.g. interleaved COSY (A‐A COSY and X‐X COSY); (b) parallel acquisition experiments, e.g.…”

“…[4.5] In this manuscript we focus on experiments involving simultaneous detection of two abundant and highly sensitive nuclei, 1 H and 19 F. Many of the experiments discussed in the succeeding texts can be (and in many Figure 1. Examples of the three types of experiments involving two receivers: (a) interleaved experiments, [28] e.g. interleaved COSY (A-A COSY and X-X COSY); (b) parallel acquisition experiments, [2] e.g.…”

“…The interleaved experiments (Fig. b–d) also known as Unified Time‐Optimized Interleaved Acquisition (UTOPIA) typically involve two or more ‘independent’ spin systems where the individual experiments are designed in such a way as to avoid mutual interferences. In a typical interleaved experiment magnetization of the passive nuclei is essentially undisturbed during the experiment involving the active nuclei and the time saving is achieved by allowing passive nuclear species A to recover while the experiment involving active nuclei X is carried out and vice versa (Fig.…”