title: Following Metabolism in Living Microorganisms by Hyperpolarized (1)H NMR.
abstract: Dissolution dynamic nuclear polarization (dDNP) is used to enhance the sensitivity of nuclear magnetic resonance (NMR), enabling monitoring of metabolism and specific enzymatic reactions in vivo. dDNP involves rapid sample dissolution and transfer to a spectrometer/scanner for subsequent signal detection. So far, most biologically oriented dDNP studies have relied on hyperpolarizing long-lived nuclear spin species such as (13)C in small molecules. While advantages could also arise from observing hyperpolarized (1)H, short relaxation times limit the utility of prepolarizing this sensitive but fast relaxing nucleus. Recently, it has been reported that (1)H NMR peaks in solution-phase experiments could be hyperpolarized by spontaneous magnetization transfers from bound (13)C nuclei following dDNP. This work demonstrates the potential of this sensitivity-enhancing approach to probe the enzymatic process that could not be suitably resolved by (13)C dDNP MR. Here we measured, in microorganisms, the action of pyruvate decarboxylase (PDC) and pyruvate formate lyase (PFL)-enzymes that catalyze the decarboxylation of pyruvate to form acetaldehyde and formate, respectively. While (13)C NMR did not possess the resolution to distinguish the starting pyruvate precursor from the carbonyl resonances in the resulting products, these processes could be monitored by (1)H NMR at 500 MHz. These observations were possible in both yeast and bacteria in minute-long kinetic measurements where the hyperpolarized (13)C enhanced, via (13)C → (1)H cross-relaxation, the signals of protons binding to the (13)C over the course of enzymatic reactions. In addition to these spontaneous heteronuclear enhancement experiments, single-shot acquisitions based on J-driven (13)C → (1)H polarization transfers were also carried out. These resulted in higher signal enhancements of the (1)H resonances but were not suitable for multishot kinetic studies. The potential of these (1)H-based approaches for measurements in vivo is briefly discussed.
authors.affiliation: Klinik und Poliklinik für Nuklearmedizin, Technische Universität München , München 81675, Germany.|Cancer Research UK Cancer Institute , Cambridge CB2 0RE, United Kingdom.
authors.forename: Piotr
authors.initials: P
authors.lastname: Dzien
authors.forename: Anne
authors.initials: A
authors.lastname: Fages
authors.forename: Ghil
authors.initials: G
authors.lastname: Jona
authors.affiliation: Cancer Research UK Cancer Institute , Cambridge CB2 0RE, United Kingdom.
authors.forename: Kevin M
authors.initials: KM
authors.lastname: Brindle
authors.affiliation: Klinik und Poliklinik für Nuklearmedizin, Technische Universität München , München 81675, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Schwaiger
authors.forename: Lucio
authors.initials: L
authors.lastname: Frydman
mesh_terms: D000123:Acetyltransferases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D011767:Pyruvate Decarboxylase / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
chemical_list: D000123:Acetyltransferases; C022560:formate C-acetyltransferase; D011767:Pyruvate Decarboxylase