Lysine Acetylation Activates Mitochondrial Aconitase in the Heart

Jolyn Fernandes, Alexis Weddle, Caroline S. Kinter, Kenneth M. Humphries, Timothy Mather, Luke I. Szweda, Michael Kinter

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Abstract

High-throughput proteomics studies have identified several thousand acetylation sites on more than 1000 proteins. Mitochondrial aconitase, the Krebs cycle enzyme that converts citrate to isocitrate, has been identified in many of these reports. Acetylated mitochondrial aconitase has also been identified as a target for sirtuin 3 (SIRT3)-catalyzed deacetylation. However, the functional significance of mitochondrial aconitase acetylation has not been determined. Using in vitro strategies, mass spectrometric analyses, and an in vivo mouse model of obesity, we found a significant acetylation-dependent activation of aconitase. Isolated heart mitochondria subjected to in vitro chemical acetylation with either acetic anhydride or acetyl-coenzyme A resulted in increased aconitase activity that was reversed with SIRT3 treatment. Quantitative mass spectrometry was used to measure acetylation at 21 lysine residues and revealed significant increases with both in vitro treatments. A high-fat diet (60% of kilocalories from fat) was used as an in vivo model and also showed significantly increased mitochondrial aconitase activity without changes in protein level. The high-fat diet also produced an increased level of aconitase acetylation at multiple sites as measured by the quantitative mass spectrometry assays. Treatment of isolated mitochondria from these mice with SIRT3 abolished the high-fat diet-induced activation of aconitase and reduced acetylation. Finally, kinetic analyses found that the increase in activity was a result of increased maximal velocity, and molecular modeling suggests the potential for acetylation at K144 to perturb the tertiary structure of the enzyme. The results of this study reveal a novel activation of mitochondrial aconitase by acetylation. (Figure Presented.)

Original languageEnglish (US)
Pages (from-to)4008-4018
Number of pages11
JournalBiochemistry
Volume54
Issue number25
DOIs
StatePublished - Jun 30 2015

Fingerprint

Aconitate Hydratase
Acetylation
Lysine
Sirtuin 3
High Fat Diet
Nutrition
Fats
Mitochondria
Chemical activation
Mass spectrometry
Mass Spectrometry
Heart Mitochondria
Acetyl Coenzyme A
Molecular modeling
Citric Acid Cycle
Enzymes
Citric Acid
Proteomics
Assays
Proteins

ASJC Scopus subject areas

  • Biochemistry
  • Medicine(all)

Cite this

Fernandes, J., Weddle, A., Kinter, C. S., Humphries, K. M., Mather, T., Szweda, L. I., & Kinter, M. (2015). Lysine Acetylation Activates Mitochondrial Aconitase in the Heart. Biochemistry, 54(25), 4008-4018. https://doi.org/10.1021/acs.biochem.5b00375

Lysine Acetylation Activates Mitochondrial Aconitase in the Heart. / Fernandes, Jolyn; Weddle, Alexis; Kinter, Caroline S.; Humphries, Kenneth M.; Mather, Timothy; Szweda, Luke I.; Kinter, Michael.

In: Biochemistry, Vol. 54, No. 25, 30.06.2015, p. 4008-4018.

Research output: Contribution to journalArticle

Fernandes, J, Weddle, A, Kinter, CS, Humphries, KM, Mather, T, Szweda, LI & Kinter, M 2015, 'Lysine Acetylation Activates Mitochondrial Aconitase in the Heart', Biochemistry, vol. 54, no. 25, pp. 4008-4018. https://doi.org/10.1021/acs.biochem.5b00375
Fernandes J, Weddle A, Kinter CS, Humphries KM, Mather T, Szweda LI et al. Lysine Acetylation Activates Mitochondrial Aconitase in the Heart. Biochemistry. 2015 Jun 30;54(25):4008-4018. https://doi.org/10.1021/acs.biochem.5b00375
Fernandes, Jolyn ; Weddle, Alexis ; Kinter, Caroline S. ; Humphries, Kenneth M. ; Mather, Timothy ; Szweda, Luke I. ; Kinter, Michael. / Lysine Acetylation Activates Mitochondrial Aconitase in the Heart. In: Biochemistry. 2015 ; Vol. 54, No. 25. pp. 4008-4018.
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