TY - JOUR
T1 - Following Metabolism in Living Microorganisms by Hyperpolarized 1H NMR
AU - Dzien, Piotr
AU - Fages, Anne
AU - Jona, Ghil
AU - Brindle, Kevin M.
AU - Schwaiger, Markus
AU - Frydman, Lucio
N1 - Publisher Copyright:
© 2016 American Chemical Society.
PY - 2016/9/21
Y1 - 2016/9/21
N2 - 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 13C in small molecules. While advantages could also arise from observing hyperpolarized 1H, short relaxation times limit the utility of prepolarizing this sensitive but fast relaxing nucleus. Recently, it has been reported that 1H NMR peaks in solution-phase experiments could be hyperpolarized by spontaneous magnetization transfers from bound 13C 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 13C 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 13C 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 1H NMR at 500 MHz. These observations were possible in both yeast and bacteria in minute-long kinetic measurements where the hyperpolarized 13C enhanced, via 13C → 1H cross-relaxation, the signals of protons binding to the 13C over the course of enzymatic reactions. In addition to these spontaneous heteronuclear enhancement experiments, single-shot acquisitions based on J-driven 13C → 1H polarization transfers were also carried out. These resulted in higher signal enhancements of the 1H resonances but were not suitable for multishot kinetic studies. The potential of these 1H-based approaches for measurements in vivo is briefly discussed.
AB - 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 13C in small molecules. While advantages could also arise from observing hyperpolarized 1H, short relaxation times limit the utility of prepolarizing this sensitive but fast relaxing nucleus. Recently, it has been reported that 1H NMR peaks in solution-phase experiments could be hyperpolarized by spontaneous magnetization transfers from bound 13C 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 13C 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 13C 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 1H NMR at 500 MHz. These observations were possible in both yeast and bacteria in minute-long kinetic measurements where the hyperpolarized 13C enhanced, via 13C → 1H cross-relaxation, the signals of protons binding to the 13C over the course of enzymatic reactions. In addition to these spontaneous heteronuclear enhancement experiments, single-shot acquisitions based on J-driven 13C → 1H polarization transfers were also carried out. These resulted in higher signal enhancements of the 1H resonances but were not suitable for multishot kinetic studies. The potential of these 1H-based approaches for measurements in vivo is briefly discussed.
UR - http://www.scopus.com/inward/record.url?scp=84988672740&partnerID=8YFLogxK
U2 - 10.1021/jacs.6b07483
DO - 10.1021/jacs.6b07483
M3 - Article
C2 - 27556338
AN - SCOPUS:84988672740
SN - 0002-7863
VL - 138
SP - 12278
EP - 12286
JO - Journal of the American Chemical Society
JF - Journal of the American Chemical Society
IS - 37
ER -