# Rhodamine6G and Hœchst33342 narrow BmrA conformational spectrum for a more efficient use of ATP

## TL;DR

This paper shows how drug binding affects the structure and ATP use of a transporter protein, improving its efficiency.

## Contribution

The study reveals that drug binding narrows conformational dynamics of BmrA, enhancing ATP efficiency and transport activity.

## Key findings

- Rhodamine6G and Hœchst33342 induce cooperative ATP binding in BmrA.
- Drug binding narrows BmrA's conformational spectrum, increasing ATPase activity.
- CryoEM structures reveal structural dynamics during transporter conformational transitions.

## Abstract

Multidrug ABC transporters harness the energy of ATP binding and hydrolysis to translocate substrates out of the cell and detoxify them. While this involves a well-accepted alternating access mechanism, molecular details of this interplay are still elusive. Rhodamine6G binding on a catalytic inactive mutant of the homodimeric multidrug ABC transporter BmrA triggers a cooperative binding of ATP on the two identical nucleotide-binding-sites, otherwise michaelian. Here, we investigate this asymmetric behavior via a structural-enzymology approach, solving cryoEM structures of BmrA at defined ATP ratios, highlighting the plasticity of BmrA as it undergoes the transition from inward to outward facing conformations. Analysis of continuous heterogeneity within cryoEM data and structural dynamics, reveals that Rhodamine6G narrows the conformational spectrum explored by the nucleotide-binding domains. We observe the same behavior for the other drug Hœchst33342. Following on these findings, the effect of drug-binding showed an ATPase stimulation and a maximal transport activity of the wild-type protein at the concentration-range where the cooperative transition occurs. Altogether, these findings provide a description of the influence of drug binding on the ATP-binding sites through a change in conformational dynamics.

ABC transporters are mobile and adaptative molecular machines. Here, Gobet et al characterize how substrate-binding influences the ABC transporter BmrA and triggers cooperativity in ATP-binding, utilizing an integrative structural enzymology approach.