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Biochem J. 2017 Aug 10;474(16):2829-2839. doi: 10.1042/BCJ20170389.

Potential mechanisms linking SIRT activity and hypoxic 2-hydroxyglutarate generation: no role for direct enzyme (de)acetylation.

Author information

1
Department of Anesthesiology, University of Rochester Medical Center, Rochester, NY 14642, U.S.A.
2
Department of Neuroscience, University of Rochester Medical Center, Rochester, NY, U.S.A.
3
Department of Pharmacology and Physiology, University of Rochester Medical Center, Rochester, NY, U.S.A.
4
Department of Medicine, University of Rochester Medical Center, Rochester, NY, U.S.A.
5
Department of Biochemistry, University of Rochester Medical Center, Rochester, NY, U.S.A.
6
Department of Proteomics Core Facility, University of Rochester Medical Center, Rochester, NY, U.S.A.
7
Department of Biology, University of Rochester Medical Center, Rochester, NY, U.S.A.
8
Department of Anesthesiology, University of Rochester Medical Center, Rochester, NY 14642, U.S.A. paul_brookes@urmc.rochester.edu.

Abstract

2-Hydroxyglutarate (2-HG) is a hypoxic metabolite with potentially important epigenetic signaling roles. The mechanisms underlying 2-HG generation are poorly understood, but evidence suggests a potential regulatory role for the sirtuin family of lysine deacetylases. Thus, we hypothesized that the acetylation status of the major 2-HG-generating enzymes [lactate dehydrogenase (LDH), isocitrate dehydrogenase (IDH) and malate dehydrogenase (MDH)] may govern their 2-HG-generating activity. In vitro acetylation of these enzymes, with confirmation by western blotting, mass spectrometry, reversibility by recombinant sirtuins and an assay for global lysine occupancy, yielded no effect on 2-HG-generating activity. In addition, while elevated 2-HG in hypoxia is associated with the activation of lysine deacetylases, we found that mice lacking mitochondrial SIRT3 exhibited hyperacetylation and elevated 2-HG. These data suggest that there is no direct link between enzyme acetylation and 2-HG production. Furthermore, our observed effects of in vitro acetylation on the canonical activities of IDH, MDH and LDH appeared to contrast with previous findings wherein acetyl-mimetic lysine mutations resulted in the inhibition of these enzymes. Overall, these data suggest that a causal relationship should not be assumed between acetylation of metabolic enzymes and their activities, canonical or otherwise.

KEYWORDS:

acetylation; epigenetics; hypoxia; ischemia; sirtuins

PMID:
28673962
PMCID:
PMC5562404
DOI:
10.1042/BCJ20170389
[Indexed for MEDLINE]
Free PMC Article

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