ABCMdb

PMID: 23921386

Randak CO, Dong Q, Ver Heul AR, Elcock AH, Welsh MJ
ATP and AMP mutually influence their interaction with the ATP-binding cassette (ABC) adenylate kinase cystic fibrosis transmembrane conductance regulator (CFTR) at separate binding sites.
J Biol Chem. 2013 Sep 20;288(38):27692-701. doi: 10.1074/jbc.M113.479675. Epub 2013 Aug 6., [PubMed]

Sentences

No. Mutations Sentence Comment

94 ABCC7 p.Ala462Phe Recordings from patches containing very few channels (A462F CFTR) with up to five simultaneous channel openings were low pass-filtered at 500 Hz for analysis. Login to comment 238 ABCC7 p.Ala462Phe Berger et al. (15) found that substituting alanine at position 462 in NBD1 with phenylalanine (A462F mutation; Fig. 9B, left) abolished nucleotide interaction with ATP-binding site 1. Login to comment 239 ABCC7 p.Ser1248Phe Substituting serine at position 1248 in NBD2 with phenylalanine (S1248F mutation; Fig. 9C, left) abolished nucleotide interaction with ATP-binding site 2. Login to comment 241 ABCC7 p.Ala462Phe ABCC7 p.Ser1248Phe The A462F, but not the S1248F mutation interfered with processing and trafficking to the cell membrane (supplemental Fig. S1), and hence, the number of channels in excised membrane patches was small; therefore, we quantified channel activity as NPo. Login to comment 242 ABCC7 p.Ala462Phe We found that Ap5A reduced the NPo of A462F CFTR (Fig. 9B, middle and right). Login to comment 243 ABCC7 p.Ser1248Phe On the other hand, Ap5A had no effect on ATP-dependent current of S1248F CFTR (Fig. 9C, middle and right). Login to comment 245 ABCC7 p.Lys1250Ala Our results are consistent with the previous observations that mutations of conserved residues in the Walker A and B motifs of ATP-binding site 2, K1250A and FIGURE 8. Login to comment 261 ABCC7 p.Ala462Phe B, left, model of A462F CFTR. Login to comment 262 ABCC7 p.Ala462Phe Middle, current recording from one excised inside-out membrane patch containing at least two A462F CFTR channels perfused on cytosolic surface with ATP and Ap5A as indicated. Login to comment 268 ABCC7 p.Ala462Phe Right, NPo of A462F CFTR with 0.3 mM ATP and PKA present in the bath solution before and after adding 1 mM Ap5A. Login to comment 270 ABCC7 p.Ser1248Phe C, left, model of S1248F CFTR. Login to comment 272 ABCC7 p.Ser1248Phe Middle, current recording (100 ms averages) from an excised inside-out membrane patch containing multiple S1248F CFTR channels. ATP and Ap5A were present during the times and at the concentrations indicated by bars. Login to comment 273 ABCC7 p.Ser1248Phe ATP was added together with PKA catalytic subunit. Holding voltage was afa;80 mV. Right, S1248F CFTR Clafa; current before and after adding 1 mM Ap5A. Experiments were performed as shown in the middle panel with 0.3 mM ATP and PKA present. Login to comment 275 ABCC7 p.Lys464Ala ABCC7 p.Asp1370Asn ABCC7 p.Asp572Asn Error bars, S.E. Nucleotide Interactions with the ABC Adenylate Kinase CFTR 27698 JOURNAL OF BIOLOGICAL CHEMISTRY VOLUME 288ߦNUMBER 38ߦSEPTEMBER 20, 2013 at SEMMELWEIS UNIV OF MEDICINE on December , D1370N, abolished Ap5A inhibition of current, whereas the homologous mutations in ATP-binding site 1, K464A and D572N, did not. Login to comment 301 ABCC7 p.Ser1248Phe (a) The S1248F mutation, which interfered with the binding of ATP to site 2 (15) and abolished the inhibition of CFTR current in the presence of Ap5A in our study (Fig. 9C), also disrupted CFTR adenylate kinase activity (22). Login to comment 302 ABCC7 p.Lys1250Ala ABCC7 p.Asp1370Asn (b) Patch clamp studies showed that CFTR mutations K1250A and D1370N, located within the conserved Walker A and B motifs of ATP-binding site 2, abolished the effects of Ap5A and AMP on CFTR current. Login to comment 303 ABCC7 p.Lys464Ala ABCC7 p.Asp572Asn However, the homologous mutations in ATP-binding site 1 (K464A and D572N) did not (19). Login to comment 321 ABCC7 p.Gln1291Arg Mutation Q1291R has been found in patients with cystic fibrosis. Login to comment 322 ABCC7 p.Gly576Ala ABCC7 p.Gln1352His ABCC7 p.Gly550Arg Mutations G576A, G550R, and Q1352H have been described in patients with congenital bilateral absence of the vas deferens, a condition that affects men with cystic fibrosis but can also occur in the absence Nucleotide Interactions with the ABC Adenylate Kinase CFTR SEPTEMBER 20, 2013ߦVOLUME 288ߦNUMBER 38 JOURNAL OF BIOLOGICAL CHEMISTRY 27699 of other disease manifestations. Login to comment 329 ABCC7 p.Leu1254Ala An example is L1254A CFTR, which showed an increase in current after adding Ap5A due to a reduced channel closing rate. Login to comment