Tang Jia, Yin Hang, Qiu Jade, Tucker Matthew J, DeGrado William F, Gai Feng
Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
J Am Chem Soc. 2009 Mar 25;131(11):3816-7. doi: 10.1021/ja809007f.
Helix-helix association within a membrane environment represents one of the fundamental processes in membrane protein folding. However, studying the kinetics of such processes has been difficult because most membrane proteins are insoluble in aqueous solution. Here we present a stopped-flow fluorescence study of the membrane-interaction kinetics of a designed, water-soluble transmembrane (TM) peptide, anti-alpha(IIb), which is known to dimerize in phospholipid bilayers. We show that by using two fluorescent amino acids, tryptophan and p-cyanophenylalanine, we are able to kinetically dissect distinct phases in the peptide-membrane interaction, representing membrane binding, membrane insertion, and TM helix-helix association. Our results further show that the last process occurs on a time scale of seconds, indicating that the association of two TM helices is an intrinsically slow event.
膜环境中的螺旋-螺旋缔合是膜蛋白折叠的基本过程之一。然而，研究此类过程的动力学一直很困难，因为大多数膜蛋白在水溶液中不溶。在此，我们展示了一项关于设计的水溶性跨膜（TM）肽抗α（IIb）的膜相互作用动力学的停流荧光研究，已知该肽在磷脂双层中会二聚化。我们表明，通过使用两种荧光氨基酸，色氨酸和对氰基苯丙氨酸，我们能够从动力学上剖析肽-膜相互作用中的不同阶段，这些阶段代表膜结合、膜插入以及跨膜螺旋-螺旋缔合。我们的结果进一步表明，最后一个过程发生在秒的时间尺度上，这表明两个跨膜螺旋的缔合是一个本质上缓慢的事件。
