FLUORESCENCE-QUENCHING-RESOLVED SPECTRA OF MELITTIN IN LIPID BILAYERS
FLUORESCENCE-QUENCHING-RESOLVED SPECTRA OF MELITTIN IN LIPID BILAYERS
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DOI：
10.1016/0167-4838(90)90131-x
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发表时间：
1990-09-27
期刊：
影响因子：
--
通讯作者：
WASYLEWSKI, Z
中科院分区：
文献类型：
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作者：
KASZYCKI, P;WASYLEWSKI, Z
用荧光猝灭法研究了蜂毒蜂毒素与二肉豆蔻基磷脂酰胆碱(DMPC)单层囊泡的相互作用。在脂肽比Ri=50和高离子强度(2MNaC L)下，用荧光猝灭法研究了蜂毒蜂毒素与DMPC单层囊泡的相互作用。采用荧光猝灭分辨光谱(FQRS)法，以碘化钾为猝灭剂，在DMPC的主相变温度(TT)以上和以下，将脂质体结合蜂毒素的色氨酸发射光谱分解成多个组分。其中一个分辨光谱的最大吸收波长分别为342 nm和338 nm，与四聚体蜂毒素在溶液中的最大色氨酸发射波长(340 Nm)相似。这一光谱的特征是Stern-Volmer猝灭常数Ksv约为4M-1，它表示其色氨酸残基在溶剂中暴露于溶剂中的蜂毒素分子的比例，其程度与溶液中的四聚物种相当。另一个光谱分量，对应于色氨酸分子的猝灭剂不可及部分(Ksv=0 M-1)，其最大蓝移至15 nm，表明环境的极性降低。对于TT实验，蓝光谱成分揭示了激发-波长依赖性，可能起源于激发色氨酸分子和脂类极性头部基团之间的弛豫过程。我们得出结论，与DMPC脂质体结合的蜂毒素以两种与脂相关的形式存在：一种是与色氨酸残基接触到溶剂中，另一种是穿透膜内，色氨酸残基位于外囊脂层的磷脂极头基团附近。我们还用目前蜂毒素-双层相互作用的模型讨论了我们的数据。
The interaction of bee venom melittin with dimyristoylphosphatidylcholine (DMPC) unilamellar vesicles has been studied by means of fluorescence quenching of the single tryptophan residue of the protein, at lipid-to-peptide ratio, Ri = 50 and at high ionic strength (2 M NaCl). The method of fluorescence-quenching-resolved spectra (FQRS), applied in this study with potassium iodide as a quencher, enabled us to decompose the tryptophan emission spectrum of liposome-bound melittin into components, at temperatures above as well as below the main phase transition temperature (Tt) of DMPC. One of the two resolved spectra exhibits maximum at 342 and 338 nm for experiments above and below Tt, respectively, and is similar to the maximum of tryptophan emission found for tetrameric melittin in solution (340 nm). This spectrum is characterized by the Stern-Volmer quenching constant, Ksv, of about 4 M-1 and it represents the fraction of melittin molecules whose tryptophan residues are exposed to the solvent to a degree comparable with tetrameric species in solution. The other spectrum component, corresponding to the quencher-inaccessible fraction of tryptophan molecules (Ksv = 0 M-1) has its maximum blue-shifted up to 15 nm, indicating a decrease in polarity of the environment. For experiments about Tt, the blue spectrum component revealed the excitation-wavelenth dependence, originating probably from the relaxation process between the excited tryptophan molecules and lipid polar head groups. We conclude that melittin bound to DMPC liposomes exists in two lipid-associated forms; one, with tryptophan residues exposed to the solvent and the other, penetrating the membrane interior, with tryptophan residues located in close proximity to the phospholipid polar head groups of the outer vesicle lipid layer. We also discuss our data with current models of melittin-bilayer interactions.