Examinando por Autor "Rubio-Moscardo, F."

Mostrando 1 - 2 de 2
  • Miniatura
    Ítem
    Structural determinants of 5′,6′-epoxyeicosatrienoic acid binding to and activation of TRPV4 channel
    (Nature Publishing Group, 2017-12) Berna-Erro, A.; Izquierdo-Serra, M.; Sepúlveda, R.V.; Rubio-Moscardo, F.; Doñate-Macián, P.; Serra, S.A.; Carrillo-Garcia, J.; Perálvarez-Marín, A.; González-Nilo, F.; Fernández-Fernández, J.M.; Valverde, M.A.
    TRPV4 cation channel activation by cytochrome P450-mediated derivatives of arachidonic acid (AA), epoxyeicosatrienoic acids (EETs), constitute a major mechanisms of endothelium-derived vasodilatation. Besides, TRPV4 mechano/osmosensitivity depends on phospholipase A2 (PLA2) activation and subsequent production of AA and EETs. However, the lack of evidence for a direct interaction of EETs with TRPV4 together with claims of EET-independent mechanical activation of TRPV4 has cast doubts on the validity of this mechanism. We now report: 1) The identification of an EET-binding pocket that specifically mediates TRPV4 activation by 5′,6′-EET, AA and hypotonic cell swelling, thereby suggesting that all these stimuli shared a common structural target within the TRPV4 channel; and 2) A structural insight into the gating of TRPV4 by a natural agonist (5′,6′-EET) in which K535 plays a crucial role, as mutant TRPV4-K535A losses binding of and gating by EET, without affecting GSK1016790A, 4α-phorbol 12,13-didecanoate and heat mediated channel activation. Together, our data demonstrates that the mechano- and osmotransducing messenger EET gates TRPV4 by a direct action on a site formed by residues from the S2-S3 linker, S4 and S4-S5 linker.
  • Miniatura
    Ítem
    Structural determinants of TRPV4 inhibition and identification of new antagonists with antiviral activity
    (John Wiley and Sons Inc, 2020) Donate-Macian, P.; Duarte, Y.; Rubio-Moscardo, F.; Perez-Vilaro, G.; Canan, J.; Diez, J.; Gonzalez-Nilo, F.; Valverde, M.
    Background and Purpose: The transient receptor potential vanilloid 4 (TRPV4) cation channel participates in multiple physiological processes and is also at the core of different diseases, making this channel an interesting pharmacological target with therapeutic potential. However, little is known about the structural elements governing its inhibition. Experimental Approach: We have now combined in silico drug discovery and molecular dynamics simulation based on Xenopus tropicalis xTRPV4 structure with functional studies measuring cell Ca2+ influx mediated by human TRPV4 channel to characterize the binding site of known TRPV4 inhibitors and to identify novel small molecule channel modulators. Key Results: We have found that the inhibitor HC067047 binds to a pocket conformed by residues from S2–S3 linker (xTRPV4-D542), S4 (xTRPV4-M583 and Y587 and S5 (xTRPV4-D609 and F613). This pocket was also used for structure-based virtual screening in the search of novel channel modulators. Forty potential hits were selected based on the lower docking scores (from ~250,000 compounds) and their effect upon TRPV4 functionally tested. Three were further analysed for stability using molecular dynamics simulation and functionally tested on TRPV4 channels carrying mutations in the binding pocket. Compound NSC151066, shown to require residue xTRPV4-M583 for its inhibitory effect, presented an IC50 of 145 nM and demonstrated to be an effective antiviral against Zika virus with a potency similar to HC067047. Conclusion and Implications: Together, we propose structural insights into the inhibition of TRPV4 and how this information can be used for the design of novel channel modulators.