Studies on the regulatory mechanism of isocitrate dehydrogenase 2 using acetylation mimics

Abstract

Mitochondrial isocitrate dehydrogenase 2 (IDH2) converts NADP+ to NADPH and promotes regeneration of reduced glutathione (GSH) by supplying NADPH to glutathione reductase or thioredoxin reductase. We have previously shown that under calorie restriction, mitochondrial deacetylase Sirt3 deacetylates and activates IDH2, thereby regulating the mitochondrial glutathione antioxidant defense system in mice. To investigate the regulatory mechanism of mIDH2 (mouse mitochondrial IDH2), we used lysine-to-glutamine (KQ) mutants to mimic acetylated lysines and screened 15 KQ mutants. Among these mutants, the activities of the K256Q and K413Q proteins were less than 50% of the wild-type value. We then solved the crystal structures of the wild-type mIDH2 and the K256Q mutant proteins, revealing conformational changes in the substrate-binding pocket. Structural data suggested that positively charged Lys256 was important in stabilizing the pocket because it repelled a lysine cluster on the other side. Glutamine (or acetylated lysine) was neutral and thus caused the pocket size to decrease, which might be the main reason for the lower activity of the K256Q mutant. Together, our data provide the first structure of an acetylation mimic of mIDH2 and new insights into the regulatory mechanism of acetylation of mIDH2.

P300/CBP‐associated factor regulates transcription and function of isocitrate dehydrogenase 2 during muscle differentiation

“…Among them, IDH2 is involved in the oxidation and decarboxylation of isocitrate to aKG (29). Specifically, IDH2 activity is sensitive to acetylation (22,27). The deacetylation of K413, in fact, increases IDH2 activity, whereas the constitutive acetylation mimic mutant, K256 to glutamine (K256 → Q), reduces it (22,27,47).…”

“…Specifically, IDH2 activity is sensitive to acetylation (22,27). The deacetylation of K413, in fact, increases IDH2 activity, whereas the constitutive acetylation mimic mutant, K256 to glutamine (K256 → Q), reduces it (22,27,47). Although proteomic analyses and site-specific mutagenesis indicated the presence of additional putatively acetylated lysines (27,48), no information is currently available about their functional role, and no evidence has been provided so far about the acetylase responsible for IDH2 modification.…”

“…The isocitrate dehydrogenase 2 (IDH2) is one of the mitochondrial metabolic enzymes acetylated at different lysine residues (22,27). This PTM negatively regulates the enzyme activity (22,27).…”

“…The isocitrate dehydrogenase 2 (IDH2) is one of the mitochondrial metabolic enzymes acetylated at different lysine residues (22,27). This PTM negatively regulates the enzyme activity (22,27). At present, 3 isoforms of IDHs have been identified: IDH1, -2, and -3, with the latter evolutionary distinct from the others (28,29).…”

“…In fact, in a reduced energy environment, such as in the presence of caloric restriction, SIRT3 deacetylates IDH2 at lysine residue 413, localized in the NADP + -binding pocket, and consequently, IDH2 is activated (22). Furthermore, acetylation of K256 has been reported to reduce IDH2 enzymatic activity, acting on the stabilization of its catalytic pocket (27).…”