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Biochim Biophys Acta Biomembr. 2017 Mar;1859(3):484-492. doi: 10.1016/j.bbamem.2017.01.006. Epub 2017 Jan 7.

Influence of glutamic acid residues and pH on the properties of transmembrane helices.

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Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, AR 72701, United States.
Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, AR 72701, United States. Electronic address:


Negatively charged side chains are important for the function of particular ion channels and certain other membrane proteins. To investigate the influence of single glutamic acid side chains on helices that span lipid-bilayer membranes, we have employed GWALP23 (acetyl-GGALW5LALALALALALALW19LAGA-amide) as a favorable host peptide framework. We substituted individual Leu residues with Glu residues (L12E or L14E or L16E) and incorporated specific 2H-labeled alanine residues within the core helical region or near the ends of the sequence. Solid-state 2H NMR spectra reveal little change for the core labels in GWALP23-E12, -E14 and -E16 over a pH range of 4 to 12.5, with the spectra being broader for samples in DOPC compared to DLPC bilayers. The spectra for samples with deuterium labels near the helix ends on alanines 3 and 21 show modest pH-dependent changes in the extent of unwinding of the helix terminals in DLPC and DOPC bilayers. The combined results indicate minor overall responses of these transmembrane helices to changes in pH, with the most buried residue E12 showing no pH dependence. While the Glu residues E14 and E16 may have high pKa values in the lipid bilayer environment, it is also possible that a paucity of helix response is masking the pKa values. Interestingly, when E16 is present, spectral changes at high pH report significant local unwinding of the core helix. Our results are consistent with the expectation that buried carboxyl groups aggressively hold their protons and/or waters of hydration.


Deuterium solid-state NMR; GWALP23 transmembrane helix; Glutamic acid titration; Helix terminal unwinding; Membrane proteins

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