Hinderliter A K, Dibble A R, Biltonen R L, Sando J J
Department of Pharmacology, University of Virginia Health Science Center, Charlottesville 22908, USA.
Biochemistry. 1997 May 20;36(20):6141-8. doi: 10.1021/bi962715d.
To test the hypothesis that activation of protein kinase C (PKC) is related to the interface between coexisting diacylglycerol- (DAG-) enriched and DAG-poor phases, the thermotropic phase behavior of the ternary mixtures dimyristoylphosphatidylcholine (DMPC)/dimyristoylphosphatidylserine (DMPS)/dioleoylglycerol (DO), DMPC/DMPS/1-palmitoyl-2-oleoylglycerol (PO), and DMPC/DMPS/dimyristoylglycerol (DM) was analyzed and compared with the ability of the lipid mixtures to support PKC activity. Differential scanning calorimetry (DSC) was used to monitor the gel-to-liquid crystalline phase transition as a function of the mole fraction of DO (chiDO), PO (chiPO), or DM (chiDM) in DMPC/DMPS (1:1) multilamellar vesicles (MLVs) and of chiDO in large unilamellar vesicles (LUVs). The addition of DAG at low mole fractions gave rise to the appearance of two or more overlapping transitions. The phase boundaries of the ternary mixtures deduced from the partial phase diagrams were chiDO = approximately 0.10 and approximately 0.3 for DMPC/DMPS/DO, chiPO = approximately 0.05 and approximately 0.4 for DMPC/DMPS/PO, and chiDM = approximately 0.025 and approximately 0.5-0.6 for DMPC/ DMPS/DM. Above these mole fractions of DAG, the transitions again became very sharp. The ability of the lipid mixtures to support activity of PKC alpha and PKC eta was examined below and above the gel-to-liquid crystalline phase transition. In the gel phase, PKC activity went through a maximum as a function of increasing mole fraction of each DAG and was restricted to lipid compositions in which coexisting phases were observed. Maximal activity decreased with increasing saturation of the DAG. In the fluid state, maximal PKC activity was shifted to higher DO mole fractions and the peak was much broader. Collectively, these data support a role for both the presence and nature of interface between compositionally distinct domains in activation of PKC.
为了验证蛋白激酶C（PKC）的激活与富含二酰基甘油（DAG）和贫DAG相之间的界面有关这一假设，分析了三元混合物二肉豆蔻酰磷脂酰胆碱（DMPC）/二肉豆蔻酰磷脂酰丝氨酸（DMPS）/二油酰甘油（DO）、DMPC/DMPS/1-棕榈酰-2-油酰甘油（PO）和DMPC/DMPS/二肉豆蔻酰甘油（DM）的热致相行为，并将其与脂质混合物支持PKC活性的能力进行了比较。差示扫描量热法（DSC）用于监测DMPC/DMPS（1:1）多层囊泡（MLV）中DO（χDO）、PO（χPO）或DM（χDM）的摩尔分数以及大单层囊泡（LUV）中χDO作为函数时的凝胶-液晶相变。在低摩尔分数下添加DAG会导致出现两个或更多重叠的转变。从部分相图推导的三元混合物的相边界对于DMPC/DMPS/DO为χDO≈0.10和约0.3，对于DMPC/DMPS/PO为χPO≈0.05和约0.4，对于DMPC/DMPS/DM为χDM≈0.025和约0.5 - 0.6。高于这些DAG的摩尔分数，转变再次变得非常尖锐。在凝胶-液晶相变之下和之上，研究了脂质混合物支持PKCα和PKCη活性的能力。在凝胶相中，PKC活性随着每种DAG摩尔分数的增加经历一个最大值，并且限于观察到共存相的脂质组成。最大活性随着DAG饱和度的增加而降低。在流体状态下，最大PKC活性转移到更高的DO摩尔分数，并且峰值更宽。总体而言，这些数据支持在PKC激活中组成不同结构域之间界面的存在和性质都发挥作用。
