Bimodal Activation of Acetyl-CoA Carboxylase by Glutamate

Adrienne N Boone,Andy Chan,Jerzy E Kulpa,Roger W Brownsey

doi:10.1074/jbc.275.15.10819

Adrienne N Boone, Andy Chan + Show 2 more

Open Access

https://doi.org/10.1074/jbc.275.15.10819

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Abstract

Acetyl-CoA carboxylase (ACC) catalyzes the formation of malonyl-CoA, an essential substrate for fatty acid biosynthesis and a potent inhibitor of fatty acid oxidation. Here, we provide evidence that glutamate may be a physiologically relevant activator of ACC. Glutamate induced the activation of both major isoforms of ACC, prepared from rat liver, heart, or white adipose tissue. In agreement with previous studies, a type 2A protein phosphatase contributed to the effects of glutamate on ACC. However, the protein phosphatase inhibitor microcystin LR did not abolish the effects of glutamate on ACC activity. Moreover, glutamate directly activated purified preparations of ACC when protein phosphatase activity was excluded. Phosphatase-independent ACC activation by glutamate was also reflected by polymerization of the enzyme as judged by size-exclusion chromatography. The sensitivity of ACC to direct activation by glutamate was diminished by treatment in vitro with AMP-activated protein kinase or cAMP-dependent protein kinase or by beta-adrenergic stimulation of intact adipose tissue. We conclude that glutamate, an abundant intracellular amino acid, induces ACC activation through complementary actions as a phosphatase activator and as a direct allosteric ligand for dephosphorylated ACC. This study supports the general hypothesis that amino acids fulfill important roles as signal molecules as well as intermediates in carbon and nitrogen metabolism.

Highlights

Acetyl-CoA carboxylase (ACC1; EC 6.4.1.2) catalyzes the ATP- and biotin-dependent formation of malonyl-CoA, an essential substrate for fatty-acid synthase and for fatty acyl chain elongation systems [1,2,3,4]
In view of the quantitative importance of glutamine in amino acid metabolism and the abundance of intracellular glutamate in many cells [35], we examined the possibility that ACC control might be sensitive to glutamate in tissues other
ACC isolated from heart and white adipose tissue by ammonium sulfate precipitation was activated upon incubation with glutamate (Fig. 1, B–D, circles; and Table I, part A)

Summary

EXPERIMENTAL PROCEDURES

Materials—Laboratory chemicals, biochemicals, and solvents were obtained as described previously [32]. ACC was preincubated (20 min, 37 °C) in pH 7.2 buffer containing MOPS (20 mM), bovine serum albumin (2 mg/ml), and EDTA (0.5 mM). Purification of Protein Kinases—AMPK (from rat liver) and PKA (from beef heart) were purified and assayed as described previously [32, 45, 46]. To induce ACC dephosphorylation, the enzyme was purified by ammonium sulfate precipitation, resuspended in the initial homogenization buffer plus NaCl (150 mM), and incubated at 37 °C for 90 min to facilitate the actions of endogenous phosphatases. ACC was purified from rat liver by ammonium sulfate precipitation and centrifuged just prior to chromatography (315,000 ϫ g, 15 min, 4 °C). Samples (250 ␮l) were subjected to chromatography at room temperature (10 ml/h); 1-ml fractions were collected; and ACC activity was determined

RESULTS

Kinetic properties of ACC

Effects of Glutamate Depend on the Extent of ACC Phosphorylation

Glutamate Promotes Polymerization of ACC

DISCUSSION