ABCMdb

PMID: 16246032

Vergani P, Basso C, Mense M, Nairn AC, Gadsby DC
Control of the CFTR channel's gates.
Biochem Soc Trans. 2005 Nov;33(Pt 5):1003-7., [PubMed]

Sentences

No. Mutations Sentence Comment

39 ABCC7 p.Lys464Ala ABCC7 p.Asp1370Asn To determine which of the two composite sites in the NBD1-NBD2 dimer is involved in channel opening, we introduced mutations at residues seen to interact directly with the bound nucleotide in the solved crystal structures, in the head of either NBD1 [K464A (Lys464 → Ala)] or NBD2 (D1370N). Login to comment 52 ABCC7 p.Ser1248Cys ABCC7 p.Ser549Cys We first studied the functional consequence of thiol-specific cross-linking in CFTR channels containing cysteine residues introduced in the NBD1 tail (S549C) and in the NBD2 head (S1248C), in a background similar to that used for the biochemical Figure 2 Statistical coupling analysis detects co-evolution between two positions corresponding to CFTR`s Arg555 (putative hydrogen bond donor) and Thr1246 (putative hydrogen bond acceptor) (A) Side-chain distribution at acceptor position in total multiple sequence alignment (histogram on left) and in each of the two subsets of alignments obtained by 'fixing` the side chain at the donor site, either to an Arg (centre), or to a Lys residue (right). Login to comment 62 ABCC7 p.Arg555Lys ABCC7 p.Arg555Lys ABCC7 p.Thr1246Asn ABCC7 p.Thr1246Asn The WT, the two single mutants (in our case R555K and T1246N) and the double mutant (R555K/T1246N) form the corners of a thermodynamic cycle (Figure 3, inset). Login to comment 68 ABCC7 p.Arg555Lys ABCC7 p.Thr1246Asn The R555K mutation did not significantly affect the apparent affinity, while the T1246N mutation reduced it to the same degree whether the residue at position 555 was Arg or Lys. Login to comment 69 ABCC7 p.Arg555Lys The effects of the Thr to Asn mutation in WT and mutant (R555K) background are thus similar, yielding a negligible energetic coupling between the two target residues ( Gint(unbound-bound) = 0.3 ± 0.5kT). Login to comment 78 ABCC7 p.Thr1246Asn ABCC7 p.Lys1250Arg Introducing the T1246N mutation in a non-hydrolytic background (mutated at a crucial lysine, K1250R [39]) strongly destabilized the open state with respect to the 3 Mutant cycle analysis using activation free energies for the opening reaction Coupling between Arg555 and Thr1246 increases as the channel approaches the open state. Login to comment 79 ABCC7 p.Arg555Lys ABCC7 p.Arg555Lys ABCC7 p.Thr1246Asn ABCC7 p.Thr1246Asn (A) Representative records from WT, single mutants R555K and T1246N, and double mutant R555K/T1246N, activated with 300 nM cAMP-dependent protein kinase and 5 mM ATP. Login to comment 83 ABCC7 p.Arg555Lys The same mutation, however, when introduced in the non-hydrolytic background that also contained the R555K mutation, did not significantly alter the closed- to-open equilibrium. Login to comment