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Controlling association of vesicle embedded peptides by alteration of the physical state of the lipid matrix

机译:通过改变脂质基质的物理状态来控制囊泡内嵌肽的缔合

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We report here the reversible association of a designed peptide embedded in a lipid membrane through a stimulus-sensitive trigger that changes the physical state of the bilayer matrix. A peptide designed with the classical 4-3 heptad repeat of coiled coils, equipped with leucine residues at all canonical interface positions, TH1, was rendered membrane soluble by replacement of all exterior residues with randomly selected hydrophobic amino acids. Insertion of TH1 into large unilamellar phosphatidylcholine vesicles was followed by monitoring tryptophan fluorescence. Peptide insertion was observed when the lipids were in the liquid-crystalline state [1-palmitoyl-2-oleoyl-siz-glycero-3-phosphocholine (POPC)] but not when they were in the crystalline phase [1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC)]. Formation of a trimeric a-helical bundle in lipid bilayers was followed by fluorescence resonance energy transfer. Global fit analysis revealed a monomer-trimer equilibrium with a dissociation constant of around 10(-6) MF2. A lipid mixture composed of DPPC and POPC exhibiting a phase transition at 34 degrees C between a crystalline/liquid-crystalline coexistence region and a completely miscible liquid-crystalline phase was used to control the formation of the trimeric peptide bundle. TH1 is phase excluded in crystalline DPPC domains below 34 degrees C, leading to a larger number of trimers. However, when the DPPC domains are dispersed at temperatures above 34 degrees C, the number of trimers is reduced.
机译:我们在这里报告通过改变双层基质的物理状态的刺激敏感触发器嵌入脂质膜中的设计肽的可逆关联。通过在所有规范界面位置TH1均配备亮氨酸残基的经典4-3螺旋卷曲螺旋重复序列设计的肽,可通过用随机选择的疏水性氨基酸置换所有外部残基,使膜可溶。将TH1插入大的单层磷脂酰胆碱囊泡中,然后监测色氨酸荧光。当脂质处于液晶状态[1-棕榈酰基-2-油酰基-siz-甘油-3-磷酸胆碱(POPC)]时观察到了肽插入,但当它们处于结晶相[1,2-二棕榈酰基- Sn-甘油-3-磷酸胆碱(DPPC)]。在脂质双层中形成三聚体α-螺旋束,然后进行荧光共振能量转移。整体拟合分析显示单体三聚体平衡,其解离常数约为10(-6)MF2。由DPPC和POPC组成的脂质混合物在34℃在晶体/液晶共存区域和完全可混溶的液晶相之间表现出相变,用于控制三聚体肽束的形成。在低于34摄氏度的结晶DPPC域中,TH1被排除在相外,导致大量的三聚体。但是,当DPPC域在高于34摄氏度的温度下分散时,三聚体的数量会减少。

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