ABCMdb

PMID: 10342826

Ashfield R, Gribble FM, Ashcroft SJ, Ashcroft FM
Identification of the high-affinity tolbutamide site on the SUR1 subunit of the K(ATP) channel.
Diabetes. 1999 Jun;48(6):1341-7., [PubMed]

Sentences

No. Mutations Sentence Comment

4 ABCC8 p.Ser1237Tyr High-affinity tolbutamide inhibition could be conferred on SUR2A by replacing transmembrane domains (TMs) 14-16 with the corresponding region of SUR1. Conversely, high-affinity tolbutamide inhibition of SUR1 was abolished by replacing TMs 13-16 with the corresponding SUR2A sequence, or by mutating a single serine residue within this region to tyrosine (S1237Y). Login to comment 32 ABCC8 p.Ser1237Tyr alent region of SUR2A or by a mutation within this region (S1237Y). Login to comment 127 ABCC8 p.Ser1201Cys ABCC8 p.Leu1226Met ABCC8 p.Ala1204Ser ABCC8 p.Thr1130Ile ABCC8 p.Cys1174Phe ABCC8 p.Tyr1229Leu ABCC8 p.Cys1128Thr ABCC8 p.Gln1223Lys ABCC8 p.Tyr1218His ABCC8 p.Cys1141Ser ABCC8 p.Gln1190Glu Most mutations did not affect tolbutamide sensitivity: these included C1128T, T1130I, C1141S, C1174F, Q1190E, S1201C, A1204S, Y1218H, Q1223K, Y1229L, L1226M (data not shown; n = 2-4 patches in each case). Login to comment 138 ABCC8 p.Ser1237Tyr While MgADP did not promote tolbutamide inhibition of Kir6.2-SUR1(S1237Y) currents, the nucleotide was able to enhance meglitinideblock(Fig. 4). Login to comment 145 ABCC8 p.Ser1237Tyr The dashed lines indicate the data for Kir6.2-SUR1 and Kir6.2-SUR2A given in A and B. D: Kir6.2-SUR1(S1237Y) currents were fit with equation 4: Ki1 = 1.3 mmol/l, h1 = .9, L = 0 (n = 5). Login to comment 153 ABCC8 p.Ser1237Tyr We comparedthe binding of [3 H]glibenclamide to SUR1, SUR12-e, and SUR1(S1237Y) expressed in Cos7 cells, and evaluatedprotein expression by immunoblotting with the M2 anti-FLAG antibody (Fig. 6). Login to comment 154 ABCC8 p.Ser1237Tyr Binding of [3 H]glibenclamide to SUR12-e or SUR1(S1237Y) was not significantly greater than that found inuntransfected cells. Login to comment 158 ABCC8 p.Ser1237Tyr High-affinity tolbutamide inhibition could be conferred on SUR2A by replacing TMs 14-16 with the equivalent region of SUR1. Conversely, the reverse chimera, or a mutation (S1237Y) within this region of SUR1, abolished both high-affinity tolbutamide block and [3 H]glibenclamide binding. Login to comment 169 ABCC8 p.Ser1237Tyr Effects of tolbutamide (A) or meglitinide (B) in the presence or absence of 100 µmol/l MgADP, for channels comprising Kir6.2 and either SUR1, SUR2A, or SUR1(S1237Y). Login to comment 179 ABCC8 p.Ser1237Tyr Oocytes were coinjected with mRNAs encoding Kir6.2 and either SUR1, SUR2A, SUR12-e, SUR21-x, or SUR1(S1237Y). Login to comment 194 ABCC8 p.Ser1237Tyr This can be most simply explained if the S1237Y mutation specifically impaired tolbutamide binding, without interfering with either meglitinide binding or the mechanism by which sulfonylureas interfere with channel activation by MgADP. Login to comment 198 ABCC8 p.Ser1237Tyr Thus, when SUR1 was rendered insensitive to tolbutamide [SUR12-e or SUR1(S1237Y)], glibenclamide block became reversible, while endowment of SUR2A with tolbutamide sensitivity (SUR21-x) was accompanied by irreversible glibenclamide inhibition. Login to comment 206 ABCC8 p.Ser1237Tyr A: Membranes from Cos7 cells expressing SUR1, SUR12-e, or SUR1(S1237Y) were separated by SDS polyacrylamide gel electrophoresis, and probed with the anti-FLAG monoclonal antibody M2. Login to comment 208 ABCC8 p.Ser1237Tyr B: [3 H]glibenclamide binding to membranes from Cos7 cells transfected with SUR1, SUR12-e, or SUR1(S1237Y) or to untransfected cells. Login to comment 220 ABCC8 p.Ser1237Tyr In support of this view, we found that SUR12-e and SUR1(S1237Y), which form KATP channels that are blocked reversibly by glibenclamide, did not exhibit significant [3 H]glibenclamide binding. Login to comment 224 ABCC8 p.Ser1237Tyr Conversely, high-affinity tolbutamide block and [3 H]glibenclamide binding were abolished by the reverse chimera or by mutation of a single amino acid, S1237Y, in the intracellular loop between TMs 15 and 16 of SUR1. Login to comment