Redox Regulation of Brain Selective Kinases BRSK1/2: Implications for Dynamic Control of the Eukaryotic AMPK family through Cys-based mechanisms

Bendzunas, G.; Byrne, D. P.; Shrestha, S.; Daly, L. A.; Oswald, S. O.; Katiyar, S.; Venkat, A.; Yeung, W.; Eyers, C. E.; Eyers, P. A.; Kannan, N.

doi:10.1101/2023.10.05.561145

bioRxiv · 10.1101/2023.10.05.561145

Redox Regulation of Brain Selective Kinases BRSK1/2: Implications for Dynamic Control of the Eukaryotic AMPK family through Cys-based mechanisms

Bendzunas, G.·Byrne, D. P.·Shrestha, S.·Daly, L. A.·Oswald, S. O.·Katiyar, S.·Venkat, A.·Yeung, W.·Eyers, C. E.·Eyers, P. A.·Kannan, N.

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Abstract

In eukaryotes, protein kinase signaling is regulated by a diverse array of post-translational modifications (PTMs), including phosphorylation of Ser/Thr residues and oxidation of cysteine (Cys) residues. While regulation by activation segment phosphorylation of Ser/Thr residues is well understood, relatively little is known about how oxidation of cysteine residues modulate catalysis. In this study, we investigate redox regulation of the AMPK-related Brain-selective kinases (BRSK) 1 and 2, and detail how broad catalytic activity is directly regulated through reversible oxidation and reduction of evolutionarily conserved Cys residues within the catalytic domain. We show that redox-dependent control of BRSKs is a dynamic and multilayered process involving oxidative modifications of several Cys residues, including the formation of intra-molecular disulfide bonds involving a pair of Cys residues near the catalytic HRD motif and a highly conserved T-Loop Cys with a BRSK-specific Cys within an unusual CPE motif at the end of the activation segment. Consistently, mutation of the CPE-Cys increases catalytic activity in vitro and drives phosphorylation of the BRSK substrate Tau in cells. Molecular modeling and molecular dynamics simulations indicate that oxidation of the CPE-Cys destabilizes a conserved salt bridge network critical for allosteric activation. The occurrence of spatially proximal Cys amino acids in diverse Ser/Thr protein kinase families suggests that disulfide mediated control of catalytic activity may be a prevalent mechanism for regulation within the broader AMPK family.

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Bendzunas, G., Byrne, D. P., Shrestha, S., Daly, L. A., Oswald, S. O., Katiyar, S., Venkat, A., Yeung, W., Eyers, C. E., Eyers, P. A., Kannan, N.. 2023-10-06. Redox Regulation of Brain Selective Kinases BRSK1/2: Implications for Dynamic Control of the Eukaryotic AMPK family through Cys-based mechanisms. https://doi.org/10.1101/2023.10.05.561145

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