Steric Interactions of Valines 1, 5, and 7 in [Valine 5, d-Alanine 8] Gramicidin A Channels

AbstractWhen the central valine residues 6, 7, and 8 of gramicidin A (gA) are shifted by one position, the resulting [Val5, d-Ala8]gA forms right-handed channels with a single-channel conductance and average duration somewhat less than gA channels. The reduction in channel duration has been attributed to steric conflict between the side chains of Val1 and Val5 in opposing monomers (Koeppe, R. E. II, D. V. Greathouse, A. Jude, G. Saberwal, L. L. Providence, and O. S. Andersen. 1994. J. Biol. Chem. 269:12567–12576). To investigate the orientations and motions of valines in [Val5, d-Ala8]gA, we have incorporated 2H labels at Val 1, 5, or 7 and recorded 2H-NMR spectra of oriented and nonoriented samples in hydrated dimyristoylphosphatidylcholine. Spectra of nonoriented samples at 4°C reveal powder patterns that indicate rapid side chain “hopping” for Val5, and an intermediate rate of hopping for Val1 and Val7 that is somewhat slower than in gA. Oriented samples of deuterated Val1 and Val7 show large changes in the methyl and Cβ-2H quadrupolar splittings (Δνq) when Ala5 in native gA is changed to Val5. Three or more peaks for the Val1 methyls with Δνq values that vary with the echo delay, together with an intermediate spectrum for nonoriented samples at 4°C, suggest unusual side chain dynamics for Val1 in [Val5, d-Ala8]gA. These results are consistent with a steric conflict that has been introduced between the two opposing monomers. In contrast, the acylation of gA has little influence on the side chain dynamics of Val1, regardless of the identity of residue 5