Journal of Biological Chemistry
Volume 279, Issue 40, 1 October 2004, Pages 41664-41669
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Membrane Transport, Structure, Function, and Biogenesis
A Heteromeric Complex of the Two Nucleotide Binding Domains of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Mediates ATPase Activity*

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The cystic fibrosis transmembrane conductance regulator (CFTR) protein is a member of the ABC superfamily of transporter proteins. Recently, crystal structures of intact, prokaryotic members of this family have been described. These structures suggested that ATP binding and hydrolysis occurs at two sites formed at the interface between their nucleotide binding domains (NBDs). In contrast to the prokaryotic family members, the NBDs of CFTR are asymmetric (both structurally and functionally), and previous to the present studies, it was not clear whether both NBDs are required for ATP hydrolysis. In order to assess the relative roles of the two NBDs of human CFTR, we purified and reconstituted NBD1 and NBD2, separately and together. We found that NBD1 and NBD2 by themselves exhibited relatively low ATPase activity. Co-assembly of NBD1 and NBD2 exhibited a 2–3-fold enhancement in catalytic activity relative to the isolated domains and this increase reflected enhanced ATP turnover (Vmax). These data provide the first direct evidence that heterodimerization of the NBD1 and NBD2 domains of CFTR is required to generate optimal catalytic activity.

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*

This work was supported by the Canadian Cystic Fibrosis Foundation through an operating grant (to C. E. B.) and a fellowship award (to J. F. K.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.