Dataset Information

Measurement of lipogenic flux by deuterium resolved mass spectrometry.

ABSTRACT: De novo lipogenesis (DNL) is disrupted in a wide range of human disease. Thus, quantification of DNL may provide insight into mechanisms and guide interventions if it can be performed rapidly and noninvasively. DNL flux is commonly measured by ²H incorporation into fatty acids following deuterated water (²H₂O) administration. However, the sensitivity of this approach is limited by the natural abundance of ¹³C, which masks detection of ²H by mass spectrometry. Here we report that high-resolution Orbitrap gas-chromatography mass-spectrometry resolves ²H and ¹³C fatty acid mass isotopomers, allowing DNL to be quantified using lower ²H₂O doses and shorter experimental periods than previously possible. Serial measurements over 24-hrs in mice detects the nocturnal activation of DNL and matches a ³H-water method in mice with genetic activation of DNL. Most importantly, DNL is detected in overnight-fasted humans in less than an hour and is responsive to feeding during a 4-h study. Thus, ²H specific MS provides the ability to study DNL in settings that are currently impractical.

SUBMITTER: Fu X

PROVIDER: S-EPMC8213799 | biostudies-literature | 2021 Jun

REPOSITORIES: biostudies-literature

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Measurement of lipogenic flux by deuterium resolved mass spectrometry.

Fu Xiaorong X Deja Stanisław S Fletcher Justin A JA Anderson Norma N NN Mizerska Monika M Vale Gonçalo G Browning Jeffrey D JD Horton Jay D JD McDonald Jeffrey G JG Mitsche Matthew A MA Burgess Shawn C SC

Nature communications 20210618 1

De novo lipogenesis (DNL) is disrupted in a wide range of human disease. Thus, quantification of DNL may provide insight into mechanisms and guide interventions if it can be performed rapidly and noninvasively. DNL flux is commonly measured by 2H incorporation into fatty acids following deuterated water (2H2O) administration. However, the sensitivity of this approach is limited by the natural abundance of 13C, which masks detection of 2H by ...[more]

PMID: 34145255

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Project description:Hydrogen/deuterium exchange mass spectrometry (HDX-MS) is a powerful technique to interrogate protein structure and dynamics. With the ability to study almost any protein without a size limit, including intrinsically disordered ones, HDX-MS has shown fast growing importance as a complement to structural elucidation techniques. Current experiments compare two or more related conditions (sequences, interaction partners, excipients, conformational states, etc.) to determine statistically significant differences at a number of fixed time points and highlight areas of changed structural dynamics in the protein. The work presented here builds on the fundamental research performed in the early days of the technique and re-examines exchange rate calculations with the aim of establishing HDX-MS as an absolute and quantitative, rather than relative and qualitative, measurement. We performed millisecond HDX-MS experiments on a mixture of three unstructured peptides (angiotensin, bradykinin, and atrial natriuretic peptide amide rat) and compared experimental deuterium uptake curves with theoretical ones predicted using established exchange rate calculations. With poly-dl-alanine (PDLA) commonly used as a reference,, we find that experimental rates are sometimes faster than theoretically possible, while they agree much better, and are never faster, with the fully unstructured trialanine peptide (3-Ala). Molecular dynamics (MD) simulations confirm the high helical propensity of the longer and partially structured PDLA peptides, which need as few as 15 residues to form a stable helix and are therefore not suitable as an unstructured reference. Reanalysis of previously published data by Weis et al. at 100 mM NaCl however still shows a discrepancy with predictions based on 3-Ala in the absence of salt, highlighting the need for a better understanding of salt effects on exchange rates. Such currently unquantifiable salt effects prevent us from proposing a comprehensive, universal calibration framework at the moment. Nevertheless, an accurate recalibration of intrinsic exchange rate calculations is crucial to enable kinetic modeling of the exchange process and to ultimately allow HDX-MS to move toward a direct link with atomistic structural models.

Dataset Information

Measurement of lipogenic flux by deuterium resolved mass spectrometry.

Publications

Measurement of lipogenic flux by deuterium resolved mass spectrometry.

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