Durkin J T, Providence L L, Koeppe R E, Andersen O S
Department of Physiology and Biophysics, Cornell University Medical College, New York, New York 10021.
Biophys J. 1992 Apr;62(1):145-57; discussion 157-9. doi: 10.1016/S0006-3495(92)81801-5.
Using the linear gramicidins as an example, we have previously shown how the statistical properties of heterodimeric (hybrid) channels (formed between the parent [Val1]gramicidin A (gA) and a sequence-altered analogue) can be used to assess whether the analogue forms channels that are structurally equivalent to the parent channels (Durkin, J. T., R. E. Koeppe II, and O. S. Andersen. 1990. J. Mol. Biol. 211:221-234). Generally, the gramicidins are tolerant of amino acid sequence alterations. We report here an exception. The optically reversed analogue, gramicidin M- (gM-) (Heitz, F., G. Spach, and Y. Trudelle. 1982. Biophys. J. 40:87-89), forms channels that are the mirror-image of [Val1]gA channels; gM- should thus form no hybrid channels with analogues having the same helix sense as [Val1]gA. Surprisingly, however, gM- forms hybrid channels with the shortened analogues des-Val1-[Ala2]gA and des-Val1-gC, but these channels differ fundamentally from the parent channels: (a) the appearance rate of these heterodimers is only approximately 1/10 of that predicted from the random assortment of monomers into conducting dimers, indicating the existence of an energy barrier to their formation (e.g., monomer refolding into a new channel-forming conformation); and (b), once formed, the hybrid channels are stabilized approximately 1,000-fold relative to the parent channels. The increased stability suggests a structure that is joined by many hydrogen bonds, such as one of the double-stranded helical dimers shown to be adopted by gramicidins in organic solvents (Veatch, W. R., E. T. Fossel, and E. R. Blout. 1974. Biochemistry. 13:5249-5256).
以线性短杆菌肽为例，我们之前已经展示了如何利用异二聚体（杂合）通道（由亲本[Val1]短杆菌肽A（gA）和序列改变的类似物之间形成）的统计特性来评估该类似物形成的通道在结构上是否等同于亲本通道（Durkin, J. T., R. E. Koeppe II, and O. S. Andersen. 1990. J. Mol. Biol. 211:221 - 234）。一般来说，短杆菌肽对氨基酸序列改变具有耐受性。我们在此报告一个例外情况。光学反转类似物，短杆菌肽M -（gM -）（Heitz, F., G. Spach, and Y. Trudelle. 1982. Biophys. J. 40:87 - 89），形成的通道是[Val1]gA通道的镜像；因此，gM - 不应与具有与[Val1]gA相同螺旋方向的类似物形成杂合通道。然而，令人惊讶的是，gM - 与缩短的类似物去-Val1-[Ala2]gA和去-Val1 - gC形成杂合通道，但这些通道与亲本通道有根本差异：（a）这些异二聚体的出现率仅约为从单体随机组合形成传导性二聚体所预测值的1/10，表明其形成存在能量障碍（例如，单体重新折叠成新的通道形成构象）；以及（b）一旦形成，杂合通道相对于亲本通道稳定约1000倍。稳定性增加表明其结构由许多氢键连接，例如短杆菌肽在有机溶剂中显示采用的双链螺旋二聚体结构之一（Veatch, W. R., E. T. Fossel, and E. R. Blout. 1974. Biochemistry. 13:5249 - 5256）。
