MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supply

Isoprenoids are a very large and diverse family of metabolites required by all living organisms. All isoprenoids derive from the double-bond isomers isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), which are produced by the methylerythritol 4-phosphate (MEP) pathway in bacteria a...

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Autores: Di, Xueni|||0000-0003-2511-0313, Ortega-Alarcon, David|||0000-0003-1885-4365, Kakumanu, Ramu|||0000-0003-4984-7766, Iglesias-Fernandez, Javier|||0000-0001-7773-2945, Diaz, Lucia|||0000-0001-8850-2571, Baidoo, Edward E. K.|||0000-0001-5787-1219, Velázquez-Campoy, Adrián|||0000-0001-5702-4538, Rodríguez Concepción, Manuel|||0000-0002-1280-2305, Pérez-Gil, Jordi|||0000-0002-5632-9556
Tipo de recurso: artículo
Fecha de publicación:2023
País:España
Institución:Universitat Autònoma de Barcelona
Repositorio:Dipòsit Digital de Documents de la UAB
Idioma:inglés
OAI Identifier:oai:ddd.uab.cat:286219
Acceso en línea:https://ddd.uab.cat/record/286219
https://dx.doi.org/urn:doi:10.1016/j.xplc.2022.100512
Access Level:acceso abierto
Palabra clave:Isoprenoids
DXS
Feedback regulation
Allosteric
Monomerization
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spelling MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supplyDi, Xueni|||0000-0003-2511-0313Ortega-Alarcon, David|||0000-0003-1885-4365Kakumanu, Ramu|||0000-0003-4984-7766Iglesias-Fernandez, Javier|||0000-0001-7773-2945Diaz, Lucia|||0000-0001-8850-2571Baidoo, Edward E. K.|||0000-0001-5787-1219Velázquez-Campoy, Adrián|||0000-0001-5702-4538Rodríguez Concepción, Manuel|||0000-0002-1280-2305Pérez-Gil, Jordi|||0000-0002-5632-9556IsoprenoidsDXSFeedback regulationAllostericMonomerizationIsoprenoids are a very large and diverse family of metabolites required by all living organisms. All isoprenoids derive from the double-bond isomers isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), which are produced by the methylerythritol 4-phosphate (MEP) pathway in bacteria and plant plastids. It has been reported that IPP and DMAPP feedback-regulate the activity of deoxyxylulose 5-phosphate synthase (DXS), a dimeric enzyme that catalyzes the main flux-controlling step of the MEP pathway. Here we provide experimental insights into the underlying mechanism. Isothermal titration calorimetry and dynamic light scattering approaches showed that IPP and DMAPP can allosterically bind to DXS in vitro, causing a size shift. In silico ligand binding site analysis and docking calculations identified a potential allosteric site in the contact region between the two monomers of the active DXS dimer. Modulation of IPP and DMAPP contents in vivo followed by immunoblot analyses confirmed that high IPP/DMAPP levels resulted in monomerization and eventual aggregation of the enzyme in bacterial and plant cells. Loss of the enzymatically active dimeric conformation allows a fast and reversible reduction of DXS activity in response to a sudden increase or decrease in IPP/DMAPP supply, whereas aggregation and subsequent removal of monomers that would otherwise be available for dimerization appears to be a more drastic response in the case of persistent IPP/DMAPP overabundance (e.g., by a blockage in their conversion to downstream isoprenoids). Our results represent an important step toward understanding the regulation of the MEP pathway and rational design of biotechnological endeavors aimed at increasing isoprenoid contents in microbial and plant systems. The MEP pathway synthesizes the universal precursors of isoprenoids in plant plastids and most bacteria. This study shows that these precursors feedback-regulate their own production by direct allosteric binding to the first enzyme of the pathway, a dimeric enzyme that becomes monomeric and eventually aggregates when levels of MEP pathway products increase. 22023-01-0120232023-01-01Articlehttp://purl.org/coar/resource_type/c_6501VoRhttp://purl.org/coar/version/c_970fb48d4fbd8a85info:eu-repo/semantics/articleapplication/pdfhttps://ddd.uab.cat/record/286219https://dx.doi.org/urn:doi:10.1016/j.xplc.2022.100512reponame:Dipòsit Digital de Documents de la UABinstname:Universitat Autònoma de BarcelonaInglésengAgencia Estatal de Investigación https://doi.org/10.13039/501100011033 PID2020-115810GB-I00Agencia Estatal de Investigación https://doi.org/10.13039/501100011033 PCI2021-121941Agencia Estatal de Investigación https://doi.org/10.13039/501100011033 BFU2016-78232-PEuropean Commission https://doi.org/10.13039/501100000780 627639Agencia Estatal de Investigación https://doi.org/10.13039/501100011033 BES-2017-080739open accesshttp://purl.org/coar/access_right/c_abf2Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, i la comunicació pública de l'obra, sempre que no sigui amb finalitats comercials, i sempre que es reconegui l'autoria de l'obra original. No es permet la creació d'obres derivades.https://creativecommons.org/licenses/by-nc-nd/4.0/info:eu-repo/semantics/openAccessoai:ddd.uab.cat:2862192026-06-06T12:50:31Z
dc.title.none.fl_str_mv MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supply
title MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supply
spellingShingle MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supply
Di, Xueni|||0000-0003-2511-0313
Isoprenoids
DXS
Feedback regulation
Allosteric
Monomerization
title_short MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supply
title_full MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supply
title_fullStr MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supply
title_full_unstemmed MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supply
title_sort MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supply
dc.creator.none.fl_str_mv Di, Xueni|||0000-0003-2511-0313
Ortega-Alarcon, David|||0000-0003-1885-4365
Kakumanu, Ramu|||0000-0003-4984-7766
Iglesias-Fernandez, Javier|||0000-0001-7773-2945
Diaz, Lucia|||0000-0001-8850-2571
Baidoo, Edward E. K.|||0000-0001-5787-1219
Velázquez-Campoy, Adrián|||0000-0001-5702-4538
Rodríguez Concepción, Manuel|||0000-0002-1280-2305
Pérez-Gil, Jordi|||0000-0002-5632-9556
author Di, Xueni|||0000-0003-2511-0313
author_facet Di, Xueni|||0000-0003-2511-0313
Ortega-Alarcon, David|||0000-0003-1885-4365
Kakumanu, Ramu|||0000-0003-4984-7766
Iglesias-Fernandez, Javier|||0000-0001-7773-2945
Diaz, Lucia|||0000-0001-8850-2571
Baidoo, Edward E. K.|||0000-0001-5787-1219
Velázquez-Campoy, Adrián|||0000-0001-5702-4538
Rodríguez Concepción, Manuel|||0000-0002-1280-2305
Pérez-Gil, Jordi|||0000-0002-5632-9556
author_role author
author2 Ortega-Alarcon, David|||0000-0003-1885-4365
Kakumanu, Ramu|||0000-0003-4984-7766
Iglesias-Fernandez, Javier|||0000-0001-7773-2945
Diaz, Lucia|||0000-0001-8850-2571
Baidoo, Edward E. K.|||0000-0001-5787-1219
Velázquez-Campoy, Adrián|||0000-0001-5702-4538
Rodríguez Concepción, Manuel|||0000-0002-1280-2305
Pérez-Gil, Jordi|||0000-0002-5632-9556
author2_role author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv Isoprenoids
DXS
Feedback regulation
Allosteric
Monomerization
topic Isoprenoids
DXS
Feedback regulation
Allosteric
Monomerization
description Isoprenoids are a very large and diverse family of metabolites required by all living organisms. All isoprenoids derive from the double-bond isomers isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), which are produced by the methylerythritol 4-phosphate (MEP) pathway in bacteria and plant plastids. It has been reported that IPP and DMAPP feedback-regulate the activity of deoxyxylulose 5-phosphate synthase (DXS), a dimeric enzyme that catalyzes the main flux-controlling step of the MEP pathway. Here we provide experimental insights into the underlying mechanism. Isothermal titration calorimetry and dynamic light scattering approaches showed that IPP and DMAPP can allosterically bind to DXS in vitro, causing a size shift. In silico ligand binding site analysis and docking calculations identified a potential allosteric site in the contact region between the two monomers of the active DXS dimer. Modulation of IPP and DMAPP contents in vivo followed by immunoblot analyses confirmed that high IPP/DMAPP levels resulted in monomerization and eventual aggregation of the enzyme in bacterial and plant cells. Loss of the enzymatically active dimeric conformation allows a fast and reversible reduction of DXS activity in response to a sudden increase or decrease in IPP/DMAPP supply, whereas aggregation and subsequent removal of monomers that would otherwise be available for dimerization appears to be a more drastic response in the case of persistent IPP/DMAPP overabundance (e.g., by a blockage in their conversion to downstream isoprenoids). Our results represent an important step toward understanding the regulation of the MEP pathway and rational design of biotechnological endeavors aimed at increasing isoprenoid contents in microbial and plant systems. The MEP pathway synthesizes the universal precursors of isoprenoids in plant plastids and most bacteria. This study shows that these precursors feedback-regulate their own production by direct allosteric binding to the first enzyme of the pathway, a dimeric enzyme that becomes monomeric and eventually aggregates when levels of MEP pathway products increase.
publishDate 2023
dc.date.none.fl_str_mv 2
2023-01-01
2023
2023-01-01
dc.type.none.fl_str_mv Article
http://purl.org/coar/resource_type/c_6501
VoR
http://purl.org/coar/version/c_970fb48d4fbd8a85
dc.type.openaire.fl_str_mv info:eu-repo/semantics/article
format article
dc.identifier.none.fl_str_mv https://ddd.uab.cat/record/286219
https://dx.doi.org/urn:doi:10.1016/j.xplc.2022.100512
url https://ddd.uab.cat/record/286219
https://dx.doi.org/urn:doi:10.1016/j.xplc.2022.100512
dc.language.none.fl_str_mv Inglés
eng
language_invalid_str_mv Inglés
language eng
dc.relation.none.fl_str_mv Agencia Estatal de Investigación https://doi.org/10.13039/501100011033 PID2020-115810GB-I00
Agencia Estatal de Investigación https://doi.org/10.13039/501100011033 PCI2021-121941
Agencia Estatal de Investigación https://doi.org/10.13039/501100011033 BFU2016-78232-P
European Commission https://doi.org/10.13039/501100000780 627639
Agencia Estatal de Investigación https://doi.org/10.13039/501100011033 BES-2017-080739
dc.rights.none.fl_str_mv open access
http://purl.org/coar/access_right/c_abf2
https://creativecommons.org/licenses/by-nc-nd/4.0/
dc.rights.openaire.fl_str_mv info:eu-repo/semantics/openAccess
rights_invalid_str_mv open access
http://purl.org/coar/access_right/c_abf2
https://creativecommons.org/licenses/by-nc-nd/4.0/
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.source.none.fl_str_mv reponame:Dipòsit Digital de Documents de la UAB
instname:Universitat Autònoma de Barcelona
instname_str Universitat Autònoma de Barcelona
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