ABCC7 p.Glu267Lys
| Predicted by SNAP2: | A: N (61%), C: D (63%), D: N (78%), F: D (80%), G: D (59%), H: D (75%), I: D (63%), K: D (59%), L: D (63%), M: D (53%), N: N (57%), P: D (71%), Q: N (66%), R: D (66%), S: N (66%), T: N (61%), V: D (53%), W: D (85%), Y: D (75%), |
| Predicted by PROVEAN: | A: N, C: D, D: N, F: D, G: N, H: N, I: D, K: N, L: D, M: D, N: N, P: N, Q: N, R: N, S: N, T: N, V: D, W: D, Y: D, |
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[hide] Biological and structural basis for Aha1 regulatio... Mol Biol Cell. 2010 Mar 15;21(6):871-84. Epub 2010 Jan 20. Koulov AV, Lapointe P, Lu B, Razvi A, Coppinger J, Dong MQ, Matteson J, Laister R, Arrowsmith C, Yates JR 3rd, Balch WE
Biological and structural basis for Aha1 regulation of Hsp90 ATPase activity in maintaining proteostasis in the human disease cystic fibrosis.
Mol Biol Cell. 2010 Mar 15;21(6):871-84. Epub 2010 Jan 20., 2010-03-15 [PMID:20089831]
Abstract [show]
The activator of Hsp90 ATPase 1, Aha1, has been shown to participate in the Hsp90 chaperone cycle by stimulating the low intrinsic ATPase activity of Hsp90. To elucidate the structural basis for ATPase stimulation of human Hsp90 by human Aha1, we have developed novel mass spectrometry approaches that demonstrate that the N- and C-terminal domains of Aha1 cooperatively bind across the dimer interface of Hsp90 to modulate the ATP hydrolysis cycle and client activity in vivo. Mutations in both the N- and C-terminal domains of Aha1 impair its ability to bind Hsp90 and stimulate its ATPase activity in vitro and impair in vivo the ability of the Hsp90 system to modulate the folding and trafficking of wild-type and variant (DeltaF508) cystic fibrosis transmembrane conductance regulator (CFTR) responsible for the inherited disease cystic fibrosis (CF). We now propose a general model for the role of Aha1 in the Hsp90 ATPase cycle in proteostasis whereby Aha1 regulates the dwell time of Hsp90 with client. We suggest that Aha1 activity integrates chaperone function with client folding energetics by modulating ATPase sensitive N-terminal dimer structural transitions, thereby protecting transient folding intermediates in vivo that could contribute to protein misfolding systems disorders such as CF when destabilized.
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No. Sentence Comment
283 Notably, these included C207S, E221A, E267K, D293A, E297A, T298A, and E313A, residues that may contribute to the stability of the Aha1 interaction with Hsp90 (Figures 3 and 4B).
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ABCC7 p.Glu267Lys 20089831:283:38
status: NEW[hide] An electrostatic interaction at the tetrahelix bun... J Biol Chem. 2014 Oct 31;289(44):30364-78. doi: 10.1074/jbc.M114.595710. Epub 2014 Sep 4. Wang W, Roessler BC, Kirk KL
An electrostatic interaction at the tetrahelix bundle promotes phosphorylation-dependent cystic fibrosis transmembrane conductance regulator (CFTR) channel opening.
J Biol Chem. 2014 Oct 31;289(44):30364-78. doi: 10.1074/jbc.M114.595710. Epub 2014 Sep 4., [PMID:25190805]
Abstract [show]
The CFTR channel is an essential mediator of electrolyte transport across epithelial tissues. CFTR opening is promoted by ATP binding and dimerization of its two nucleotide binding domains (NBDs). Phosphorylation of its R domain (e.g. by PKA) is also required for channel activity. The CFTR structure is unsolved but homology models of the CFTR closed and open states have been produced based on the crystal structures of evolutionarily related ABC transporters. These models predict the formation of a tetrahelix bundle of intracellular loops (ICLs) during channel opening. Here we provide evidence that residues E267 in ICL2 and K1060 in ICL4 electrostatically interact at the interface of this predicted bundle to promote CFTR opening. Mutations or a thiol modifier that introduced like charges at these two positions substantially inhibited ATP-dependent channel opening. ATP-dependent activity was rescued by introducing a second site gain of function (GOF) mutation that was previously shown to promote ATP-dependent and ATP-independent opening (K978C). Conversely, the ATP-independent activity of the K978C GOF mutant was inhibited by charge- reversal mutations at positions 267 or 1060 either in the presence or absence of NBD2. The latter result indicates that this electrostatic interaction also promotes unliganded channel opening in the absence of ATP binding and NBD dimerization. Charge-reversal mutations at either position markedly reduced the PKA sensitivity of channel activation implying strong allosteric coupling between bundle formation and R domain phosphorylation. These findings support important roles of the tetrahelix bundle and the E267-K1060 electrostatic interaction in phosphorylation-dependent CFTR gating.
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No. Sentence Comment
96 The charge-reversal mutations (E267R and K1060E) at these positions clearly had the most dramatic effects on CFTR activity (the E267K mutation also markedly inhibited CFTR activity; see Fig. 5).
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ABCC7 p.Glu267Lys 25190805:96:128
status: NEW138 Fig. 5, A--C confirm this prediction for an E267K/K1060E double mutant.
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ABCC7 p.Glu267Lys 25190805:138:44
status: NEW177 Conversely introducing a charge swap double mutation (E267K/K1060E) had no apparent effect on the basal currents and potentiator responses of this truncation construct (Fig. 7, C-E).
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ABCC7 p.Glu267Lys 25190805:177:54
status: NEW205 Importantly, normal PKA sensitivity was restored by introducing charge swap mutations across the putative interface (E267K/K1060E; Fig. 9, D-F).
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ABCC7 p.Glu267Lys 25190805:205:117
status: NEW241 A, macroscopic current record for a double mutant with charge-reversal substitutions at both positions (E267K/K1060E-CFTR).
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ABCC7 p.Glu267Lys 25190805:241:104
status: NEW247 Note that the E267K single mutation strongly inhibited channel activity as did the E267R mutation shown in Fig. 2.
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ABCC7 p.Glu267Lys 25190805:247:14
status: NEW325 Symbols are means afe; S.E. Ns are 5, 5, 4, and 5 for WT (black symbols), E267R (red), K1060E (blue), and E267K/K1060E (green), respectively.
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ABCC7 p.Glu267Lys 25190805:325:109
status: NEW