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Unravelling the Allosteric Targeting of PHGDH at the ACT-Binding Domain with a Photoactivatable Diazirine Probe and Mass Spectrometry Experiments

Authors :
Quentin Spillier
Séverine Ravez
Simon Dochain
Didier Vertommen
Léopold Thabault
Olivier Feron
Raphaël Frédérick
Source :
Molecules, Vol 26, Iss 2, p 477 (2021)
Publication Year :
2021
Publisher :
MDPI AG, 2021.

Abstract

The serine biosynthetic pathway is a key element contributing to tumor proliferation. In recent years, targeting of phosphoglycerate dehydrogenase (PHGDH), the first enzyme of this pathway, intensified and revealed to be a promising strategy to develop new anticancer drugs. Among attractive PHGDH inhibitors are the α-ketothioamides. In previous work, we have demonstrated their efficacy in the inhibition of PHGDH in vitro and in cellulo. However, the precise site of action of this series, which would help the rational design of new inhibitors, remained undefined. In the present study, the detailed mechanism-of-action of a representative α-ketothioamide inhibitor is reported using several complementary experimental techniques. Strikingly, our work led to the identification of an allosteric site on PHGDH that can be targeted for drug development. Using mass spectrometry experiments and an original α-ketothioamide diazirine-based photoaffinity probe, we identified the 523Q-533F sequence on the ACT regulatory domain of PHGDH as the binding site of α-ketothioamides. Mutagenesis experiments further documented the specificity of our compound at this allosteric site. Our results thus pave the way for the development of new anticancer drugs using a completely novel mechanism-of-action.

Details

Language :
English
ISSN :
14203049
Volume :
26
Issue :
2
Database :
Directory of Open Access Journals
Journal :
Molecules
Publication Type :
Academic Journal
Accession number :
edsdoj.397fdc8286c44ecab44d04acdbe41aea
Document Type :
article
Full Text :
https://doi.org/10.3390/molecules26020477