Diurnal regulation of the function of the rat brain glutamate dehydrogenase by acetylation and its dependence on thiamine administration

Abstract

Glutamate dehydrogenase (GDH) is essential for the brain function and highly regulated, according to its role in metabolism of the major excitatory neurotransmitter glutamate. Here we show a diurnal pattern of the GDH acetylation in rat brain, associated with specific regulation of GDH function. Mornings the acetylation levels of K84 (near the ADP site), K187 (near the active site), and K503 (GTP‐binding) are highly correlated. Evenings the acetylation levels of K187 and K503 decrease, and the correlations disappear. These daily variations in the acetylation adjust the GDH responses to the enzyme regulators. The adjustment is changed when the acetylation of K187 and K503 shows no diurnal variations, as in the rats after a high dose of thiamine. The regulation of GDH function by acetylation is confirmed in a model system, where incubation of the rat brain GDH with acetyl‐CoA changes the enzyme responses to GTP and ADP, decreasing the activity at subsaturating concentrations of substrates. Thus, the GDH acetylation may support cerebral homeostasis, stabilizing the enzyme function during diurnal oscillations of the brain metabolome. Daytime and thiamine interact upon the (de)acetylation of GDH in vitro. Evenings the acetylation of GDH from control animals increases both IC50GTP and EC50ADP. Mornings the acetylation of GDH from thiamine‐treated animals increases the enzyme IC50GTP. Molecular mechanisms of the GDH regulation by acetylation of specific residues are proposed. For the first time, diurnal and thiamine‐dependent changes in the allosteric regulation of the brain GDH due to the enzyme acetylation are shown.

Severe Spinal Cord Injury in Rats Induces Chronic Changes in the Spinal Cord and Cerebral Cortex Metabolism, Adjusted by Thiamine That Improves Locomotor Performance

“…The overview of experimental procedures and all animal groups involved in the study is presented in Figure 1. The randomization procedure employed GraphPad QuickCalcs webservice 1 as described earlier (Aleshin et al, 2020). The blinding was based on the fact that different experimenters performed separate steps of the study.…”

“…The quantification employed nano-LC-MS/MS analysis on a hybrid dual-pressure linear ion trap/orbitrap mass spectrometer (LTQ Orbitrap Velos Pro, Thermo Scientific, San Jose, United States) equipped with an U3000 nano-flow HPLC (Thermo Scientific, San Jose, CA) as described previously (Aleshin et al, 2020). Briefly, the samples were obtained after the in-gel trypsinolysis of the gel areas corresponding to GDH.…”

“…These peptides were also used to estimate the GDH expression. The comparison of the GDH protein level between different samples employed normalization to the total ion current during the mass-spectrometric analysis of the samples (Aleshin et al, 2020).…”

“…A decrease in the OGDHC activity is known to be addressed by increased thiamine influx into the affected neurons and brain (Mkrtchyan et al, 2016). Moreover, the non-coenzyme binding of thiamine or its derivatives also regulates other enzymes of the considered pathways (Mkrtchyan et al, 2015;Tsepkova et al, 2017;Aleshin et al, 2019;Aleshin et al, 2020;Mezhenska et al, 2020). Complex effects of the thiamine administration on the TABLE 1 | Metabolic profiling of free amino acids and related compounds in the brain cortex of the rats subjected to laminectomy (LE) or severe spinal cord injury (SCI), non-treated or after administration of thiamine or TEGP.…”

“…LE animals (Figure 3B). In view of the recently characterized thiamine effects on the GDH acetylation (Aleshin et al, 2020), we assessed the thiamine involvement in the regulatory acetylation of GDH in SCI animals. Figure 3D shows that in the brain of SCI animals a statistically significant increase in the relative acetylation level of the GDH residue K503 is induced by the thiamine administration.…”