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Inborn errors of metabolism (IEMs) are rare diseases produced by the accumulation of abnormal amounts of metabolites, toxic to the newborn. When not detected on time, they can lead to irreversible physiological and psychological sequels or even demise. Metabolomics has emerged as an efficient and powerful tool for IEM detection in newborns, children, and adults with late onset. In here, we screened urine samples from a large set of neonates (470 individuals) from a homogeneous population (Basque Country), for the identification of congenital metabolic diseases using NMR spectroscopy. Absolute quantification allowed to derive a probability function for up to 66 metabolites that adequately describes their normal concentration ranges in newborns from the Basque Country. The absence of another 84 metabolites, considered abnormal, was routinely verified in the healthy newborn population and confirmed for all but 2 samples, of which one showed toxic concentrations of metabolites associated to ketosis and the other one a high trimethylamine concentration that strongly suggested an episode of trimethylaminuria. Thus, a non-invasive and readily accessible urine sample contains enough information to assess the potential existence of a substantial number (>70) of IEMs in newborns, using a single, automated and standardized H-1-NMR-based analysis. The authors thank the collaboration of the Basque Biobank/BioCruces Node www.biobancovasco.org for collecting the samples and data from the neonates included in this study. Support was provided from The Department of Industry, Tourism and Trade of the Government of the Autonomous Community of the Basque Country (Elkartek BG2015 & BG2017); grant from Agencia Estatal de Investigacion (Spain) CTQ2015-68756-R and for the Severo Ochoa Excellence Accreditation (SEV-2016-0644).

In TROSY experiments, spin state selection (S3) retains only the single HSQC sub-spectrum with minimal T2 relaxation and maximal resolution, yet at the cost of eliminating half of the available polarisation as undesired anti-TROSY component. We here introduce queued TROSY (qTROSY) as a novel scheme to partially recover and exploit this anti-TROSY polarisation in two concatenated scans. After initial orthogonal spin state separation (oS3), anti-TROSY polarisation is explicitly stored while its TROSY counterpart follows the desired coherence pathway recorded in a first scan A. The immediately appended scan B then quantitatively converts the recovered anti-TROSY polarisation into a second TROSY spectrum, skipping the time-limiting long reequilibration delay. Both concatenated qTROSY scans thus ideally exploit the full initial polarisation within almost the same measurement time. In practice, T2 relaxation losses accruing during the coupling evolution delays reduced anti-TROSY polarisation recovery below 40%, obviating sensitivity enhancement through addition of both qTROSY scans; yet, scan B retained a complete scan A spectrum with up to 75% intensity. We therefore propose to employ qTROSY asymmetrically, compacting two separate conventional into one queued TROSY-type experiment with significantly reduced measurement time, implying primarily the concatenation of different three- or higher-dimensional experiments. Both anti-TROSY polarisation recoveries and possible time savings are largest for deuterated and smaller non-deuterated proteins, extending the rentability limit of the TROSY principle towards smaller molecular weights.

The backbone dynamics of the bacteriorhodopsin fragment (1–36)BR solubilized in a 1:1 chloroform/methanol mixture were investigated by heteronuclear 1H-15N NMR spectroscopy. The heteronuclear 15N longitudinal and transverse relaxation rates and 15N{1H} steady-state NOEs were measured at three magnetic fields (11.7, 14.1, and 17.6 T). Careful statistical analysis resulted in the selection of the extended model-free form of the spectral density function [Clore et al. (1990) J. Am. Chem. Soc., 112, 4989–4991] for all the backbone amides of (1–36)BR. The peptide exhibits motions on the micro-, nano-, and picosecond time scales. The dynamics of the α-helical part of the peptide (residues 9–31) are characterised by nanosecond and picosecond motions with mean order parameters S 2 = 0.60 and S 2 = 0.84, respectively. The nanosecond motions were attributed to the peptide's helix-coil transitions in equilibrium. Residues 3–7 and 30–35 also exhibit motions on the pico- and nanosecond time scales, but with lower order parameters. Residue 10 at the beginning of the α-helix and residues 30–35 at the C-terminus are involved in conformational exchange processes on the microsecond time scale. The implications of the obtained results for the studies of helix-coil transitions and the dynamics of membrane proteins are discussed.

The question is addressed of how maximal structural NOE data on double labelled proteins can be acquired with a minimal set of NOESY experiments. Two 3D-NOESY spectra are reported which, in concert with other commonly used spectra, provide a convenient strategy for NOE assignment. The 3D CNH-NOESY and 3D NCH-NOESY provide NOE connectivities between amide protons and carbon-bound protons and constitute orthogonal heteronuclear filters which eliminate diagonal signals, considerably improving spectral quality. Two different heteronuclear chemical shift dimensions are recorded in the spectra, thus exploiting the extra dispersion of the heteronucleus and considerably simplifying assignment.