A new family of protein kinases— The mitochondrial protein kinases

Protein phosphatase 2Cm is a critical regulator of branched-chain amino acid catabolism in mice and cultured cells

“…It has long been established that BCKD enzymatic activity is determined by the phosphorylation status of its E1α subunit. The BCKD kinase was purified and had its molecular identity determined almost 2 decades ago (20,21,32). However, the putative BCKD phosphatase thought to be responsible for its activation proved more elusive.…”

“…Under low levels of BCAA, BCKD is inactivated through phosphorylation at Ser293 of its E1α subunit. In the presence of BCAA and BCKA, BCKD becomes activated as Ser293 is dephosphorylated, presumably through an intramitochondrial phosphatase (18,21,22). Despite the longstanding recognition of its importance, the search in the past 2 decades for the molecular identity of the endogenous BCKD phosphatase has been unsuccessful, leaving much of the underlying mechanisms in BCAA catabolic regulation unexplored.…”

Cytosolic Branched Chain Aminotransferase (BCATc) Regulates mTORC1 Signaling and Glycolytic Metabolism in CD4+ T Cells

“…BCAT Enzyme Activity-Total branched chain aminotransferase activity was measured in solubilized cells at 37°C in 25 mM potassium phosphate buffer, pH 7.8, using 1 mM ␣-keto [1][2][3][4][5][6][7][8][9][10][11][12][13][14] C]isovalerate and 12 mM Ile as described (7). A unit of enzyme activity was defined as 1 nmol of Val formed per min/mg of protein at 37°C.…”

“…For the Leu oxidation assay, the washed T cells were incubated in Krebs buffer in the presence of 1 mM [1][2][3][4][5][6][7][8][9][10][11][12][13][14] C]Leu (specific activity 200 dpm/nmol). The assay was carried out in a 20-ml scintillation vial, and cells were gassed with a 95% O 2 , 5% CO 2 mixture for 20 s, sealed with a rubber stopper, and incubated in Dubnoff metabolic shaker at 37°C for 120 min.…”

“…BCKDC catalyzes irreversible oxidative decarboxylation of the BCKAs to produce the branched chain acyl-CoAs and NADH and contains multiple copies of three enzymes: branched chain ␣-keto acid decarboxylase/dehydrogenase (E1), dihydrolipoyl transcylase (E2), and dihydrolipoamide dehydrogenase (E3) (8). BCKDC is inactivated by phosphorylation of the E1␣ subunit of the E1 enzyme, catalyzed by a specific kinase (BDK), which is inhibited by the ␣-keto acid of Leu (KIC, ␣-ketoisocaproate) (9,10).…”