ABCC8 p.Arg74Asp
| Predicted by SNAP2: | A: N (53%), C: D (63%), D: D (80%), E: N (53%), F: D (63%), G: D (95%), H: N (82%), I: N (57%), K: N (87%), L: N (78%), M: N (57%), N: N (61%), P: N (53%), Q: N (66%), S: N (61%), T: N (61%), V: N (57%), W: D (91%), Y: D (75%), |
| Predicted by PROVEAN: | A: D, C: D, D: D, E: D, F: D, G: D, H: D, I: D, K: N, L: D, M: D, N: D, P: D, Q: D, S: D, T: D, V: D, W: D, Y: D, |
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[hide] N-terminal transmembrane domain of SUR1 controls g... J Gen Physiol. 2011 Mar;137(3):299-314. Epub 2011 Feb 14. Pratt EB, Tewson P, Bruederle CE, Skach WR, Shyng SL
N-terminal transmembrane domain of SUR1 controls gating of Kir6.2 by modulating channel sensitivity to PIP2.
J Gen Physiol. 2011 Mar;137(3):299-314. Epub 2011 Feb 14., [PMID:21321069]
Abstract [show]
Functional integrity of pancreatic adenosine triphosphate (ATP)-sensitive potassium (K(ATP)) channels depends on the interactions between the pore-forming potassium channel subunit Kir6.2 and the regulatory subunit sulfonylurea receptor 1 (SUR1). Previous studies have shown that the N-terminal transmembrane domain of SUR1 (TMD0) interacts with Kir6.2 and is sufficient to confer high intrinsic open probability (P(o)) and bursting patterns of activity observed in full-length K(ATP) channels. However, the nature of TMD0-Kir6.2 interactions that underlie gating modulation is not well understood. Using two previously described disease-causing mutations in TMD0 (R74W and E128K), we performed amino acid substitutions to study the structural roles of these residues in K(ATP) channel function in the context of full-length SUR1 as well as TMD0. Our results revealed that although R74W and E128K in full-length SUR1 both decrease surface channel expression and reduce channel sensitivity to ATP inhibition, they arrive there via distinct mechanisms. Mutation of R74 uniformly reduced TMD0 protein levels, suggesting that R74 is necessary for stability of TMD0. In contrast, E128 mutations retained TMD0 protein levels but reduced functional coupling between TMD0 and Kir6.2 in mini-K(ATP) channels formed by TMD0 and Kir6.2. Importantly, E128K full-length channels, despite having a greatly reduced P(o), exhibit little response to phosphatidylinositol 4,5-bisphosphate (PIP(2)) stimulation. This is reminiscent of Kir6.2 channel behavior in the absence of SUR1 and suggests that TMD0 controls Kir6.2 gating by modulating Kir6.2 interactions with PIP(2). Further supporting this notion, the E128W mutation in full-length channels resulted in channel inactivation that was prevented or reversed by exogenous PIP(2). These results identify a critical determinant in TMD0 that controls Kir6.2 gating by controlling channel sensitivity to PIP(2). Moreover, they uncover a novel mechanism of K(ATP) channel inactivation involving aberrant functional coupling between SUR1 and Kir6.2.
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No. Sentence Comment
100 Because of significantly reduced surface expression in most mutants, cells were pretreated with 300 µM tolbutamide overnight, which partially corrects the trafficking defect caused by R74W, as reported previously (Yan et al., 2004, 2007; Pratt et al., 2009), as well as all other R74 mutations, with the exception of R74D (Fig. S1).
X
ABCC8 p.Arg74Asp 21321069:100:321
status: NEW101 Because R74D had no detectable surface expression before or after tolbutamide exposure, it was not analyzed.
X
ABCC8 p.Arg74Asp 21321069:101:8
status: NEW263 Accordingly, with the exception of lysine, all other amino acids used in the R74X screen (A, H, C, W, F, Y, L, D, or E) are predicted to shift the boundary of TM1 and TM2 (as well as TM3 for R74D and R74E) by TOPCONS (Fig. S2).
X
ABCC8 p.Arg74Asp 21321069:263:191
status: NEW265 Sulfonylureas acting as chemical chaperones partially overcome the trafficking defect in R74X full-length KATP channels, with the exception of R74D (Fig. S1).
X
ABCC8 p.Arg74Asp 21321069:265:143
status: NEW