ABCMdb

PMID: 16223764

Zhou Z, Wang X, Li M, Sohma Y, Zou X, Hwang TC
High affinity ATP/ADP analogues as new tools for studying CFTR gating.
J Physiol. 2005 Dec 1;569(Pt 2):447-57. Epub 2005 Oct 13., 2005-12-01 [PubMed]

Sentences

No. Mutations Sentence Comment

20 ABCC7 p.Glu1371Ser By examining macroscopic and microscopic kinetics of a hydrolysis-deficient mutant CFTR (i.e. E1371S), we demonstrate an [ATP]-dependent distribution of the open time constants, indicating that ATP binding can affect the life time of the open state (Bompadre et al. 2005a,b). Login to comment 60 ABCC7 p.Glu1371Ser For E1371S-CFTR current relaxation experiments (Fig. 7A), initially channels were activated by PKA plus P-ATP in some patches, while by PKA plus ATP in others. Login to comment 63 ABCC7 p.Glu1371Ser Since we did not find time-dependent changes in E1371S-CFTR experiments or in WT-CFTR experiments, data from each set of experiments were pooled for statistical analysis. Login to comment 71 ABCC7 p.Glu1371Ser Point mutation E1371S was introduced into the pcDNA3.1 wild-type CFTR by QuikChange XL method (Stratagene, La Jolla, CA, USA). Login to comment 164 ABCC7 p.Glu1371Ser Wefurtherdifferentiatedbetweenthesetwopossibilities by using the hydrolysis-deficient mutant CFTR, E1371S (Vergani et al. 2003; Bompadre et al. 2005b). Login to comment 168 ABCC7 p.Glu1371Ser Interestingly, macroscopic E1371S-CFTR channel currents can be activated by 50 µm P-ATP and PKA, indicating that P-ATP not only can support ATP-dependent gating, but also can beusedasasubstrateforPKA-dependentphosphorylation of the R domain. Login to comment 171 ABCC7 p.Glu1371Ser These results suggest that P-ATP stabilizes the locked open state of E1371S-CFTR due to tight binding. Login to comment 189 ABCC7 p.Glu1371Ser Effect of P-ATP on CFTR locked open state A, macroscopic E1371S-CFTR current relaxations upon removal of 50 µM P-ATP or 1 mM ATP. Login to comment 191 ABCC7 p.Glu1371Ser B, mean data of the current relaxation experiments of E1371S-CFTR. Login to comment 192 ABCC7 p.Glu1371Ser The current relaxation time constants (τrelaxation) for E1371S-CFTR channels are 297.6 ± 34.0 s (n = 5) upon the removal of 50 µM P-ATP, and 118.8 ± 9.4 s (n = 5) upon the removal of 1 mM ATP. Login to comment 220 ABCC7 p.Lys464Ala Our earlier studies show that a mutation (i.e. K464A) that causes a decrease of ATP binding affinity at the NBD1 site (Basso et al. 2003) does not affect the opening rate or the ATP dose-response relationship (Powe et al. 2002; cf. Vergani et al. 2003), suggesting ATP binding at the NBD1 site may not be absolutely required for channel opening (also see Bompadre et al. 2005b). Login to comment 222 ABCC7 p.Lys464Ala The proposition that P-ATP acts on the NBD1 site to modulate channel closing is also based on our earlier studies with the K464A mutant. Login to comment 223 ABCC7 p.Lys464Ala Although this mutation does not affect channel opening, K464A-CFTR exhibits a shorter open time (Powe et al. 2002). Login to comment 224 ABCC7 p.Lys1250Ala ABCC7 p.Lys464Ala ABCC7 p.Lys464Ala ABCC7 p.Lys464Ala ABCC7 p.Glu1371Ser In addition, K464A mutation decreases the locked open time of hydrolysis-deficient mutants K464A/K1250A and K464A/E1371S (Powe et al. 2002; Vergani et al. 2003; Bompadre et al. 2005b), supporting the idea that the strength of ligand binding at the NBD1 site affects the stability of the open state. Login to comment