Biochemical evidence for ATPase activity in CFTR-enriched apical membrane vesicles from tracheal epithelium

摘要

In apical membrane vesicles from beef tracheal epithelia expressing up to 30% of the proteins as functional cystic fibrosis transmembrane conductance regulator (CFTR) - i.e. a voltage-independent and PKA-sensitive ~(36)Cl~- flux - an ATPase activity, different from P, F_0F_1 and V types, was reproducibly detected. Its specific activity averaged 20 μmol Pi h~(-1) mg~(-1) with an apparent affinity for ATP of 530 ± 30 μM. Its possible involvement in CFTR functions was supported by (1) the linear relationship between the ATPase activity and the magnitude of ~(36)Cl~- fluxes (turnover rate: 3 ATP hydrolyzed per CFTR per second), (2) the same rank of potency of ATP, ITP, GTP, UTP and CTP to be hydrolyzed and to open CFTR chloride channels, (3) the similar and parallel inhibition of the ATPase and CFTR Cl~- fluxes by NS004 (IC_(50): 60 μM) and (4) the potency of anti-R domain antibodies to increase by 18% the ATPase activity.