ABCMdb

PMID: 14697202

Randak C, Welsh MJ
An intrinsic adenylate kinase activity regulates gating of the ABC transporter CFTR.
Cell. 2003 Dec 26;115(7):837-50., [PubMed]

Sentences

No. Mutations Sentence Comment

228 ABCC7 p.Asn1303Lys ABCC7 p.Lys1250Ala ABCC7 p.Asp1370Asn In addition, we Walker A and B motif mutations (K1250A and D1370N) abolished both ATPase and adenylate kinase activities studied a relatively common CF-associated mutation, N1303K (Osborne et al., 1992); interestingly, two other (Figures 7A and 7B). Login to comment 229 ABCC7 p.Asn1303Lys In contrast, the N1303K variant only modestly impacted ATP hydrolysis: Vmax was reduced missense mutations at this site also cause CF. Login to comment 232 ABCC7 p.Asn1303Lys These data indicate that the N1303K al., 2000; Gaudet and Wiley, 2001; Hung et al., 1998; Karpowich et al., 2001; Yuan et al., 2001). Login to comment 262 ABCC7 p.Asn1303Ala We also made the corresponding mutations in CFTR of ATP, and agrees with our earlier finding that wild-type and N1303A CFTR had the same ATP EC50 for current and examined their effect on channel activity. Login to comment 263 ABCC7 p.Lys1250Ala ABCC7 p.Lys464Ala The Walker A mutations (K464A in NBD1 and K1250A in stimulation (Berger et al., 2002). Login to comment 265 ABCC7 p.Asn1303Lys These results are con- scribed above suggest that the N1303K mutation had little consequence for the ATP binding site. Login to comment 266 ABCC7 p.Asn1303Lys In contrast, sistent with the previously reported rightward shifts in the ATP dose-response curves and a reduction in ATP N1303K abolished the effects of AMP and Ap5A (Figures 7E and 7F). Login to comment 269 ABCC7 p.Lys464Ala Thus, these data begin to discriminate between the ATPase and adenylate kinase effects whereas K464A enhanced Ap5A inhibition (Figures 7E and 7F). Login to comment 272 ABCC7 p.Asp1370Asn ABCC7 p.Asp572Asn Results with the Walker B Asp Discussion mutations (D572N in NBD1 and D1370N in NBD2) exactly paralleled the P loop mutations (Figures 7D-7F), further Earlier work indicated that CFTR can function as an ATPase and that hydrolysis contributes to channel gat- suggesting that adenylate kinase activity resides in NBD2. Login to comment 274 ABCC7 p.Asn1303Lys Our data indi- We obtained different results with the N1303K mutation. Login to comment 277 ABCC7 p.Asn1303Lys This result suggests that N1303K did not alter the potency suggest that NBD2 harbors the adenylate kinase acti- Figure 7. Login to comment 280 ABCC7 p.Asn1303Lys ABCC7 p.Lys1250Ala ABCC7 p.Asp1370Asn NBD2 was wild-type or contained K1250A, D1370N, or N1303K mutations. Login to comment 283 ABCC7 p.Asn1303Lys Line for N1303K is fit using Michaelis-Menten equation with Km of 625 afe; 117 òe;M and Vmax of 6.7 afe; 0.3 nmol/(min&#b7;mg MBP-NBD2). Login to comment 288 ABCC7 p.Asn1303Lys ABCC7 p.Lys1250Ala ABCC7 p.Lys464Ala ABCC7 p.Asp1370Asn ABCC7 p.Asp572Asn Data are from 5 (wild-type, K464A, D572N), 9 (K1250A), 10 (D1370N), and 3 (N1303K) membrane patches. Asterisks indicate p b0d; 0.05 compared to wild-type by ANOVA followed by Dunnett`s multiple comparison test. Login to comment 582 ABCC7 p.Asn1303Lys On the mechanism Incidence and expression of the N1303K mutation of the cystic of MgATP-dependent gating of CFTR Clafa; channels. Login to comment