Prenner E J, Lewis R N, Jelokhani-Niaraki M, Hodges R S, McElhaney R N
Department of Biochemistry, University of Alberta, Edmonton, AB, Canada T6G 2H7.
Biochim Biophys Acta. 2001 Feb 9;1510(1-2):83-92. doi: 10.1016/s0005-2736(00)00337-0.
We have investigated the effect of the presence of 25 mol percent cholesterol on the interactions of the antimicrobial peptide gramicidin S (GS) with phosphatidylcholine and phosphatidylethanolamine model membrane systems using a variety of methods. Our circular dichroism spectroscopic measurements indicate that the incorporation of cholesterol into egg phosphatidylcholine vesicles has no significant effect on the conformation of the GS molecule but that this peptide resides in a range of intermediate polarity as compared to aqueous solution or an organic solvent. Our Fourier transform infrared spectroscopic measurements confirm these findings and demonstrate that in both cholesterol-containing and cholesterol-free dimyristoylphosphatidylcholine liquid-crystalline bilayers, GS is located in a region of intermediate polarity at the polar--nonpolar interfacial region of the lipid bilayer. However, GS appears to be located in a more polar environment nearer the bilayer surface when cholesterol is present. Our (31)P-nuclear magnetic resonance studies demonstrate that the presence of cholesterol markedly reduces the tendency of GS to induce the formation of inverted nonlamellar phases in model membranes composed of an unsaturated phosphatidylethanolamine. Finally, fluorescence dye leakage experiments indicate that cholesterol inhibits the GS-induced permeabilization of phosphatidylcholine vesicles. Thus in all respects the presence of cholesterol attenuates but does not abolish the interactions of GS with, and the characteristic effects of GS on, phospholipid bilayers. These findings may explain why it is more potent at disrupting cholesterol-free bacterial than cholesterol-containing eukaryotic membranes while nevertheless disrupting the integrity of the latter at higher peptide concentrations. This additional example of the lipid specificity of GS may aid in the rational design of GS analogs with increased antibacterial but reduced hemolytic activities.
我们使用多种方法研究了25摩尔百分比的胆固醇的存在对抗菌肽短杆菌肽S（GS）与磷脂酰胆碱和磷脂酰乙醇胺模型膜系统相互作用的影响。我们的圆二色光谱测量表明，将胆固醇掺入卵磷脂酰胆碱囊泡中对GS分子的构象没有显著影响，但与水溶液或有机溶剂相比，该肽存在于一系列中等极性的环境中。我们的傅里叶变换红外光谱测量证实了这些发现，并表明在含胆固醇和不含胆固醇的二肉豆蔻酰磷脂酰胆碱液晶双层中，GS位于脂质双层的极性-非极性界面区域的中等极性区域。然而，当存在胆固醇时，GS似乎位于更靠近双层表面的极性更强的环境中。我们的³¹P核磁共振研究表明，胆固醇的存在显著降低了GS在由不饱和磷脂酰乙醇胺组成的模型膜中诱导形成反向非层状相的倾向。最后，荧光染料泄漏实验表明，胆固醇抑制了GS诱导的磷脂酰胆碱囊泡的通透性。因此，在所有方面，胆固醇的存在都会减弱但不会消除GS与磷脂双层的相互作用以及GS对磷脂双层的特征性影响。这些发现可能解释了为什么GS在破坏不含胆固醇的细菌膜方面比含胆固醇的真核细胞膜更有效，同时在更高的肽浓度下仍能破坏后者的完整性。GS这种脂质特异性的额外例子可能有助于合理设计具有增强抗菌活性但降低溶血活性的GS类似物。
