Department of Chemistry, University of Illinois at Chicago , 845 West Taylor Street (m/c 111), Chicago, Illinois 60607-7061, United States.
Biochemistry. 2014 May 20;53(19):3079-87. doi: 10.1021/bi500027x. Epub 2014 May 12.
β-Lactoglobulin (βLG) is a member of the lipocalin protein family that changes structure upon interacting with anionic surfactants and lipid vesicles under higher-pH conditions at which βLG is dimeric. In this study, a β-sheet to α-helix transformation was also observed for monomeric βLG obtained at pH 2.6 when it was mixed with small unilamellar vesicles (SUVs) of zwitterionic lipids, but being mixed with anionic lipids produced little change. The dynamics and extent of this change were quite dependent on the lipid character, phase, and vesicle size. With 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), at ~50 °C and pH 2.6, the βLG converted to a substantially helical form upon addition of ~10 mM lipid in a two-step kinetic process having time constants of ~1 and ~25 h, as monitored by circular dichroism (CD). Fluorescence changes were simpler but implied a rapid initial change in the Trp environments followed by a slower process paralleling the change in secondary structure. Polarization attenuated total reflectance Fourier transform infrared results indicate the formed helices are at least partially inserted into the lipid bilayer and the sheet segments are on the surface. Thermal behavior showed that the secondary structure of the lipid-bound βLG had two phases, the first being characteristic of the protein-lipid vesicle interaction and the second following the DSPC phase change after which the protein apparently dissociated from the vesicle. Large unilamellar vesicles had a weaker interaction, as judged by CD, which may correlate to the partial exposure of the hydrophobic parts of the SUV bilayer. Other zwitterionic lipids bound βLG with much slower kinetics and often required sonication to induce interaction, but these also showed dissociation upon lipid phase change. These thermal and kinetic behaviors suggest a mechanism for the interaction of monomeric βLG with zwitterionic lipids different from that seen previously for the dimeric form.
β-乳球蛋白（βLG）是亲脂素蛋白家族的成员，在更高 pH 值条件下与阴离子表面活性剂和脂质体相互作用时，其结构会发生变化，此时βLG 呈二聚体形式。在本研究中，当单体βLG 在 pH 2.6 时与两性离子脂质的小单层囊泡（SUV）混合时，也观察到βLG 从β-折叠到α-螺旋的转变，但与阴离子脂质混合时几乎没有变化。这种变化的动力学和程度与脂质特性、相态和囊泡大小密切相关。对于 1,2-二硬脂酰-sn-甘油-3-磷酸胆碱（DSPC），在50°C 和 pH 2.6 下，βLG 在添加约 10 mM 脂质后，在两步动力学过程中迅速转化为主要的螺旋形式，时间常数约为1 和~25 h，如圆二色性（CD）监测所示。荧光变化较为简单，但表明色氨酸环境发生了快速初始变化，随后是与二级结构变化平行的较慢过程。偏振衰减全反射傅里叶变换红外结果表明，形成的螺旋至少部分插入脂质双层，片状片段位于表面。热行为表明，脂质结合的βLG 的二级结构有两个相，第一个是蛋白质-脂质囊泡相互作用的特征，第二个是在 DSPC 相变之后，蛋白质显然从囊泡中解离。大单层囊泡的相互作用较弱，根据 CD 判断，这可能与 SUV 双层的疏水区部分部分暴露有关。其他两性离子脂质与βLG 的结合动力学较慢，通常需要超声处理才能诱导相互作用，但这些脂质在脂质相变化时也会解离。这些热和动力学行为表明，单体βLG 与两性离子脂质的相互作用机制与先前观察到的二聚体形式不同。
