Bradrick T D, Freire E, Georghiou S
Department of Physics, University of Tennessee, Knoxville 37996-1200.
Biochim Biophys Acta. 1989 Jun 26;982(1):94-102. doi: 10.1016/0005-2736(89)90179-x.
High-sensitivity differential scanning calorimetry has been used to examine the interaction of bee venom melittin with dipalmitoylphosphatidylcholine fused unilamellar vesicles. Experiments were performed under conditions for which melittin in solution is either monomeric (in low salt) or tetrameric (in high salt). It was found that under both sets of conditions melittin abolishes the pretransition at a relatively high lipid-to-protein molar incubation ratio, Ri (about 200) and that at intermediate values of Ri it broadens the main transition profile and reduces the transition enthalpy. This provides evidence which suggests that melittin is at least partially inserted into the apolar region of the bilayer. Evident at low values of Ri are two peaks in the lipid thermal transition profiles, which may arise from a heterogeneous population of lipid vesicles formed through fusion induced by melittin, or by lipid phase separation. For those profiles which exhibited only one peak, transition enthalpies, normalized to those of the lipid in the absence of the protein, are plotted vs. the bound protein-to-lipid molar ratios for the experiments performed under the conditions which give monomeric and tetrameric melittin in solution. These plots yield straight lines, the slopes of which give the number of lipid molecules each protein molecule excludes from participating in the phase transition. These were found to be 9.9 +/- 0.7 and 4.1 +/- 0.5 for monomeric and tetrameric melittin, respectively. The results are discussed in terms of possible models for the binding of melittin to phospholipid vesicles. For simple hexagonal packing of lipid molecules, incorporation as an aggregate is favored when melittin is tetrameric in solution, whereas incorporation as a monomer is favored when melittin is monomeric in solution. For low-salt solutions, evidence is obtained for the contribution of free melittin to lipid fusion, perhaps by the formation of protein bridges between apposed vesicles.
高灵敏度差示扫描量热法已被用于研究蜂毒溶血磷脂与二棕榈酰磷脂酰胆碱融合单层囊泡的相互作用。实验是在溶液中的溶血磷脂为单体（低盐浓度）或四聚体（高盐浓度）的条件下进行的。结果发现，在这两种条件下，溶血磷脂在相对较高的脂质与蛋白质摩尔孵育比（Ri，约为200）时会消除预转变，而在Ri的中间值时，它会拓宽主要转变曲线并降低转变焓。这提供了证据表明溶血磷脂至少部分插入到双层膜的非极性区域。在Ri值较低时，脂质热转变曲线中出现两个峰，这可能是由溶血磷脂诱导融合形成的脂质囊泡异质性群体，或由脂质相分离引起的。对于那些只显示一个峰的曲线，将转变焓归一化为无蛋白质时脂质的转变焓，并针对在溶液中产生单体和四聚体溶血磷脂的条件下进行的实验，绘制其与结合的蛋白质与脂质摩尔比的关系图。这些图得出直线，其斜率给出每个蛋白质分子排除在相变中参与的脂质分子数量。对于单体和四聚体溶血磷脂，分别发现为9.9±0.7和4.1±0.5。根据溶血磷脂与磷脂囊泡结合的可能模型对结果进行了讨论。对于脂质分子的简单六方堆积，当溶液中的溶血磷脂为四聚体时，有利于以聚集体形式掺入，而当溶液中的溶血磷脂为单体时，有利于以单体形式掺入。对于低盐溶液，获得了游离溶血磷脂对脂质融合有贡献的证据，可能是通过在相邻囊泡之间形成蛋白质桥。
