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Combining Small-Molecule Bioconjugation and Hydrogen-Deuterium Exchange Mass Spectrometry (HDX-MS) to Expose Allostery: the Case of Human Cytochrome P450 3A4.

Combining Small-Molecule Bioconjugation and Hydrogen-Deuterium Exchange Mass Spectrometry (HDX-MS) to Expose Allostery: the Case of Human Cytochrome P450 3A4.

Authors :
Ducharme J
Polic V
Thibodeaux CJ
Auclair K
Source :
ACS chemical biology [ACS Chem Biol] 2021 May 21; Vol. 16 (5), pp. 882-890. Date of Electronic Publication: 2021 Apr 29.
Publication Year :
2021

Abstract

We report a novel approach to study allostery which combines the use of carefully selected bioconjugates and hydrogen-deuterium exchange mass spectrometry (HDX-MS). This strategy avoids issues related to weak substrate binding and ligand relocalization. The utility of our method is demonstrated using human cytochrome P450 3A4 (CYP3A4), the most important drug-metabolizing enzyme. Allosteric activation and inhibition of CYP3A4 by pharmaceuticals is an important mechanism of drug interactions. We performed HDX-MS analysis on several CYP3A4-effector bioconjugates, some of which mimic the allosteric effect of positive effectors, while others show activity enhancement even though the label does not occupy the allosteric pocket (agonistic) or do not show activation while still blocking the allosteric site (antagonistic). This allowed us to better define the position of the allosteric site, the protein structural dynamics associated with allosteric activation, and the presence of coexisting conformers.

Details

Language :
English
ISSN :
1554-8937
Volume :
16
Issue :
5
Database :
MEDLINE
Journal :
ACS chemical biology
Publication Type :
Academic Journal
Accession number :
33913317
Full Text :
https://doi.org/10.1021/acschembio.1c00084