Chattopadhyay Amitabha, Rawat Satinder S, Greathouse Denise V, Kelkar Devaki A, Koeppe Roger E
Centre for Cellular and Molecular Biology, Hyderabad, India.
Biophys J. 2008 Jul;95(1):166-75. doi: 10.1529/biophysj.107.124206. Epub 2008 Mar 13.
The linear peptide gramicidin forms prototypical ion channels specific for monovalent cations and has been used extensively to study the organization, dynamics, and function of membrane-spanning channels. The tryptophan residues in gramicidin channels are crucial for maintaining the structure and function of the channel. We explored the structural basis for the reduction in channel conductance in the case of single-tryptophan analogs of gramicidin with three Trp --> hydrophobic substitutions using a combination of fluorescence approaches, which include red edge excitation shift and membrane penetration depth analysis, size-exclusion chromatography, and circular dichroism spectroscopy. We show here that the gramicidin analogs containing single-tryptophan residues adopt a mixture of nonchannel and channel conformations, as evident from analysis of membrane penetration depth, size-exclusion chromatography, and backbone circular dichroism data. These results are potentially useful in analyzing the effect of tryptophan substitution on the functioning of other ion channels and membrane proteins.
线性肽短杆菌肽形成了对单价阳离子具有特异性的典型离子通道，并已被广泛用于研究跨膜通道的结构、动力学和功能。短杆菌肽通道中的色氨酸残基对于维持通道的结构和功能至关重要。我们结合荧光方法，包括红边激发位移和膜穿透深度分析、尺寸排阻色谱法和圆二色光谱法，探索了具有三个色氨酸向疏水取代的短杆菌肽单色氨酸类似物通道电导降低的结构基础。我们在此表明，含有单色氨酸残基的短杆菌肽类似物采用非通道和通道构象的混合物，这从膜穿透深度分析、尺寸排阻色谱法和主链圆二色数据中可以明显看出。这些结果对于分析色氨酸取代对其他离子通道和膜蛋白功能的影响可能是有用的。
