Mechanism of GAPDH Redox Signaling by H<sub>2</sub>O<sub>2</sub> Activation of a Two−Cysteine Switch
Oxidation of glyceraldehyde−3−phosphate dehydrogenase (GAPDH) by reactive oxygen species such as H<sub>2</sub>O<sub>2</sub> activate pleiotropic signaling pathways is associated with pathophysiological cell fate decisions. Oxidized GAPDH binds chaperone proteins with transloc...
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| Format: | Article |
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MDPI AG
2022-04-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/23/9/4604 |
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| author | Paul A. Hyslop Michael O. Chaney |
| author_facet | Paul A. Hyslop Michael O. Chaney |
| author_sort | Paul A. Hyslop |
| collection | DOAJ |
| description | Oxidation of glyceraldehyde−3−phosphate dehydrogenase (GAPDH) by reactive oxygen species such as H<sub>2</sub>O<sub>2</sub> activate pleiotropic signaling pathways is associated with pathophysiological cell fate decisions. Oxidized GAPDH binds chaperone proteins with translocation of the complex to the nucleus and mitochondria initiating autophagy and cellular apoptosis. In this study, we establish the mechanism by which H<sub>2</sub>O<sub>2</sub>−oxidized GAPDH subunits undergo a subunit conformational rearrangement. H<sub>2</sub>O<sub>2</sub> oxidizes both the catalytic cysteine and a vicinal cysteine (four residues downstream) to their respective sulfenic acids. A ‘two−cysteine switch’ is activated, whereby the sulfenic acids irreversibly condense to an intrachain thiosulfinic ester resulting in a major metastable subunit conformational rearrangement. All four subunits of the homotetramer are uniformly and independently oxidized by H<sub>2</sub>O<sub>2</sub>, and the oxidized homotetramer is stabilized at low temperatures. Over time, subunits unfold forming disulfide−linked aggregates with the catalytic cysteine oxidized to a sulfinic acid, resulting from thiosulfinic ester hydrolysis via the highly reactive thiosulfonic ester intermediate. Molecular Dynamic Simulations provide additional mechanistic insights linking GAPDH subunit oxidation with generating a putative signaling conformer. The low−temperature stability of the H<sub>2</sub>O<sub>2</sub>−oxidized subunit conformer provides an operable framework to study mechanisms associated with gain−of−function activities of oxidized GAPDH to identify novel targets for the treatment of neurodegenerative diseases. |
| first_indexed | 2024-03-10T04:06:52Z |
| format | Article |
| id | doaj.art-0afa49bb12214515ade3bfd87a843067 |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T04:06:52Z |
| publishDate | 2022-04-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-0afa49bb12214515ade3bfd87a8430672023-11-23T08:19:38ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672022-04-01239460410.3390/ijms23094604Mechanism of GAPDH Redox Signaling by H<sub>2</sub>O<sub>2</sub> Activation of a Two−Cysteine SwitchPaul A. Hyslop0Michael O. Chaney1Arkley Research Labs, Arkley BioTek, LLC, 4444 Decatur Blvd., Indianapolis, IN 46241, USAEli Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, IN 46285, USAOxidation of glyceraldehyde−3−phosphate dehydrogenase (GAPDH) by reactive oxygen species such as H<sub>2</sub>O<sub>2</sub> activate pleiotropic signaling pathways is associated with pathophysiological cell fate decisions. Oxidized GAPDH binds chaperone proteins with translocation of the complex to the nucleus and mitochondria initiating autophagy and cellular apoptosis. In this study, we establish the mechanism by which H<sub>2</sub>O<sub>2</sub>−oxidized GAPDH subunits undergo a subunit conformational rearrangement. H<sub>2</sub>O<sub>2</sub> oxidizes both the catalytic cysteine and a vicinal cysteine (four residues downstream) to their respective sulfenic acids. A ‘two−cysteine switch’ is activated, whereby the sulfenic acids irreversibly condense to an intrachain thiosulfinic ester resulting in a major metastable subunit conformational rearrangement. All four subunits of the homotetramer are uniformly and independently oxidized by H<sub>2</sub>O<sub>2</sub>, and the oxidized homotetramer is stabilized at low temperatures. Over time, subunits unfold forming disulfide−linked aggregates with the catalytic cysteine oxidized to a sulfinic acid, resulting from thiosulfinic ester hydrolysis via the highly reactive thiosulfonic ester intermediate. Molecular Dynamic Simulations provide additional mechanistic insights linking GAPDH subunit oxidation with generating a putative signaling conformer. The low−temperature stability of the H<sub>2</sub>O<sub>2</sub>−oxidized subunit conformer provides an operable framework to study mechanisms associated with gain−of−function activities of oxidized GAPDH to identify novel targets for the treatment of neurodegenerative diseases.https://www.mdpi.com/1422-0067/23/9/4604oxidative stressredox signalingglyceraldehyde−3−phosphate dehydrogenasehydrogen peroxidetwo−cysteine redox switchthiosulfinic ester |
| spellingShingle | Paul A. Hyslop Michael O. Chaney Mechanism of GAPDH Redox Signaling by H<sub>2</sub>O<sub>2</sub> Activation of a Two−Cysteine Switch International Journal of Molecular Sciences oxidative stress redox signaling glyceraldehyde−3−phosphate dehydrogenase hydrogen peroxide two−cysteine redox switch thiosulfinic ester |
| title | Mechanism of GAPDH Redox Signaling by H<sub>2</sub>O<sub>2</sub> Activation of a Two−Cysteine Switch |
| title_full | Mechanism of GAPDH Redox Signaling by H<sub>2</sub>O<sub>2</sub> Activation of a Two−Cysteine Switch |
| title_fullStr | Mechanism of GAPDH Redox Signaling by H<sub>2</sub>O<sub>2</sub> Activation of a Two−Cysteine Switch |
| title_full_unstemmed | Mechanism of GAPDH Redox Signaling by H<sub>2</sub>O<sub>2</sub> Activation of a Two−Cysteine Switch |
| title_short | Mechanism of GAPDH Redox Signaling by H<sub>2</sub>O<sub>2</sub> Activation of a Two−Cysteine Switch |
| title_sort | mechanism of gapdh redox signaling by h sub 2 sub o sub 2 sub activation of a two cysteine switch |
| topic | oxidative stress redox signaling glyceraldehyde−3−phosphate dehydrogenase hydrogen peroxide two−cysteine redox switch thiosulfinic ester |
| url | https://www.mdpi.com/1422-0067/23/9/4604 |
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