MEP pathway products allosterically promote monomerization of deoxy-D-xylulose-5-phosphate synthase to feedback-regulate their supply
Di, Xueni (Centre de Recerca en Agrigenòmica)
Ortega-Alarcon, David (Universidad de Zaragoza. Departamento de Bioquímica y Biología Molecular y Celular)
Kakumanu, Ramu (Lawrence Berkeley National Laboratory)
Iglesias-Fernandez, Javier (Nostrum Biodiscovery)
Diaz, Lucia (Nostrum Biodiscovery)
Baidoo, Edward E.K. (Lawrence Berkeley National Laboratory)
Velázquez-Campoy, Adrian (Universidad de Zaragoza. Departamento de Bioquímica y Biología Molecular y Celular)
Rodríguez Concepción, Manuel (Centre de Recerca en Agrigenòmica)
Pérez-Gil, Jordi (Centre de Recerca en Agrigenòmica)

Fecha:	2023
Resumen:	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.
Ayudas:	Agencia Estatal de Investigación PID2020-115810GB-I00 Agencia Estatal de Investigación UToPIQ-PCI2021-121941 Agencia Estatal de Investigación BFU2016-78232-P European Commission 627639 Agencia Estatal de Investigación BES-2017-080739
Derechos:	Aquest 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.
Lengua:	Anglès
Documento:	Article ; recerca ; Versió publicada
Materia:	Isoprenoids ; DXS ; Feedback regulation ; Allosteric ; Monomerization
Publicado en:	Plant Communications, Vol. 4, Issue 3 (May 2023) , art. 100512, ISSN 2590-3462

DOI: 10.1016/j.xplc.2022.100512
PMID: 36575800

12 p, 1.7 MB

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