Koeppe R E, Greathouse D V, Jude A, Saberwal G, Providence L L, Andersen O S
Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville 72701.
J Biol Chem. 1994 Apr 29;269(17):12567-76.
Gramicidin A (gA) channels are dimers formed by right-handed beta 6.3-helical monomers. The stereochemical basis for the preference of a right-handed conformation remains obscure, but it has earlier been demonstrated that the handedness can be shifted by changing the chirality of each residue in the LD-sequence and therefore is determined by the peptide itself and not by channel-membrane interactions. We now examine the contributions of Trp15, the central Val residues 6-8, and residues 1-5. None of these alone are sufficient to specify the helix sense. To examine the D-Val6-L-Val7-D-Val8 sequence, the register of the 3 valines was shifted by one to L-Val5-D-Val6-L-Val7. The resulting analogue, [Val5,D-Ala8]gA, forms channels with a conductance and duration that are both somewhat less than those of gA channels. The reduced channel duration can be attributed to a steric conflict between the side chains of Val1 in one monomer and Val5 in the other monomer. The helix handedness is not altered by this modification, as shown by circular dichroism and two-dimensional nuclear magnetic resonance spectroscopy and by hybrid channel experiments. [Val5,D-Ala8]gA forms hybrid channels with gA (which forms right-handed channels), but not with des-Val1-gA- (which forms left-handed channels). Similar hybrid channel analysis shows that des-Trp15-gA and [L-Ala1,D-Ala2,L-Ala3,D-Ala4]gA also form right-handed channels. We conclude that the helix handedness most probably is a complex function of the arrangement of both the D-Val-L-Val-D-Val and the L-Trp-(D-Leu-L-Trp)3 segments.
短杆菌肽A（gA）通道是由右手β6.3 - 螺旋单体形成的二聚体。右手构象偏好的立体化学基础仍不清楚，但早期已证明，通过改变LD序列中每个残基的手性可以改变螺旋方向，因此螺旋方向由肽本身决定，而非通道 - 膜相互作用。我们现在研究色氨酸15、中央缬氨酸残基6 - 8以及残基1 - 5的作用。这些单独一个都不足以确定螺旋方向。为研究D - 缬氨酸6 - L - 缬氨酸7 - D - 缬氨酸8序列，将3个缬氨酸的排列顺序移动一位变为L - 缬氨酸5 - D - 缬氨酸6 - L - 缬氨酸7。所得类似物[缬氨酸5，D - 丙氨酸8]gA形成的通道，其电导和持续时间均略小于gA通道。通道持续时间缩短可归因于一个单体中缬氨酸1的侧链与另一个单体中缬氨酸5的侧链之间的空间冲突。如圆二色光谱、二维核磁共振光谱以及杂交通道实验所示，这种修饰并未改变螺旋方向。[缬氨酸5，D - 丙氨酸8]gA与gA（形成右手通道）形成杂交通道，但不与去缬氨酸1 - gA（形成左手通道）形成杂交通道。类似的杂交通道分析表明，去色氨酸15 - gA和[L - 丙氨酸1，D - 丙氨酸2，L - 丙氨酸3，D - 丙氨酸4]gA也形成右手通道。我们得出结论，螺旋方向很可能是D - 缬氨酸 - L - 缬氨酸 - D - 缬氨酸和L - 色氨酸 - （D - 亮氨酸 - L - 色氨酸）3片段排列的复杂函数。
