Asymmetry of movements in CFTR's two ATP sites during pore opening serves their distinct functions

Abstract
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Abstract

CFTR, the chloride channel mutated in cystic fibrosis (CF) patients, is opened by ATP binding to two cytosolic nucleotide binding domains (NBDs), but pore-domain mutations may also impair gating. ATP-bound NBDs dimerize occluding two nucleotides at interfacial binding sites; one site hydrolyzes ATP, the other is inactive. The pore opens upon tightening, and closes upon disengagement, of the catalytic site following ATP hydrolysis. Extent, timing, and role of non-catalytic-site movements are unknown. Here we exploit equilibrium gating of a hydrolysis-deficient mutant and apply F value analysis to compare timing of opening-associated movements at multiple locations, from the cytoplasmic ATP sites to the extracellular surface. Marked asynchrony of motion in the two ATP sites reveals their distinct roles in channel gating. The results clarify the molecular mechanisms of functional cross-talk between canonical and degenerate ATP sites in asymmetric ABC proteins, and of the gating defects caused by two common CF mutations.

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Ben Sorum

Department of Medical Biochemistry, Semmelweis University, Budapest, Hungary

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0001-6742-1094
Beáta Töröcsik

Department of Medical Biochemistry, Semmelweis University, Budapest, Hungary

Competing interests
No competing interests declared.
László Csanády

Department of Medical Biochemistry, Semmelweis University, Budapest, Hungary

For correspondence
csanady.laszlo@med.semmelweis-univ.hu

Competing interests
László Csanády, Reviewing Editor, eLife.

"This ORCID iD identifies the author of this article:" 0000-0002-6547-5889

Funding

Howard Hughes Medical Institute (International Early Career Scientist Award)

László Csanády

Cystic Fibrosis Foundation (Research Grant CSANAD17G0)

László Csanády

Magyar Tudományos Akadémia (Lendület grant LP2017-14/2017)

László Csanády

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: This study was performed in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. All of the animals were handled according to approved institutional animal care and use committee (IACUC) protocols of Semmelweis University (last approved 06-30-2016, expiration 06-30-2021).

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.