An Isozyme-specific Redox Switch in Human Brain Glycogen Phosphorylase Modulates Its Allosteric Activation by AMP

Abstract

Edited by Jeffrey PessinBrain glycogen and its metabolism are increasingly recognized as major players in brain functions. Moreover, alteration of glycogen metabolism in the brain contributes to neurodegenerative processes. In the brain, both muscle and brain glycogen phosphorylase isozymes regulate glycogen mobilization. However, given their distinct regulatory features, these two isozymes could confer distinct metabolic functions of glycogen in brain. Interestingly, recent proteomics studies have identified isozymespecific reactive cysteine residues in brain glycogen phosphorylase (bGP). In this study, we show that the activity of human bGP is redox-regulated through the formation of a disulfide bond involving a highly reactive cysteine unique to the bGP isozyme. We found that this disulfide bond acts as a redox switch that precludes the allosteric activation of the enzyme by AMP without affecting its activation by phosphorylation. This unique regulatory feature of bGP sheds new light on the isoform-specific regulation of glycogen phosphorylase and glycogen metabolism.

Molecular Mechanisms of Allosteric Inhibition of Brain Glycogen Phosphorylase by Neurotoxic Dithiocarbamate Chemicals

“…To better understand the molecular mechanisms underlying the neurotoxic effects of DTCs, we investigated the potential molecular and functional effects of a neurotoxic DTC pesticide thiram, on bGP. We found that thiram (and certain of its metabolites), inhibited bGP through the formation of an intramolecular disulfide bond between Cys 318 and Cys 326 , two residues involved in the redox regulation of bGP (23). Altogether, our results suggest that the modulation of bGP activity by DTCs may participate in the neurotoxic effects of these chemicals.…”

“…Sulfoxide metabolites of DTCs are known to induce bonds (34), as well as adducts on cysteines (24). Recently, bGP has been shown to be regulated through the formation of intramolecular disulfide bonds that do not impact the migration profile of the enzyme (23). Consequently, bGP inhibition by DEDC-sulfoxide is likely to be due to either adduction of cysteine residues and/or formation of intramolecular disulfide bonds that do not impact migration profile.…”

“…Moreover, the activity of bGP can be redox-regulated through the formation of an intramolecular disulfide bond between Cys 318 and Cys 326 , which controls the AMP-dependent activation of this GP isozyme. Together, these tight regulations of bGP enzyme may participate to control of glycogen metabolism in the brain (23).…”

“…Rat models and recent proteomic studies suggested that brain glycogen phosphorylase is a putative target of DTC chemicals in the brain (7). In addition, this isoenzyme has been described as presenting highly reactive cysteine residues (23,(25)(26)(27). To further determine the impact of DTCs on bGP activity and glycogen metabolism in the brain, we treated mice brain extracts with thiram (Fig.…”

“…Consequently, bGP inhibition by DEDC-sulfoxide is likely to be due to either adduction of cysteine residues and/or formation of intramolecular disulfide bonds that do not impact migration profile. 4, a and b), the first one being specific for the brain isoform of GP (Cys 326 is not present in muscle and liver GPs) and involved in the redox regulation of bGP activity (23). To test whether this disulfide bond was involved in bGP inhibition by thiram, we performed site-directed mutagenesis experiments, substituting Cys 318 and Cys 326 with serine residues, and treated the mutated enzymes with thiram.…”