Medkova M, Cho W
Department of Chemistry (M/C 111), University of Illinois at Chicago, 845 West Taylor Street, Chicago, Illinois 60607-7061, USA.
Biochemistry. 1998 Apr 7;37(14):4892-900. doi: 10.1021/bi972495j.
To elucidate the mechanisms of membrane binding and activation of conventional and novel protein kinase C (PKC), we measured the interactions of rat PKC-alpha and -epsilon with phospholipid monolayers and vesicles of various compositions. Besides the established difference in calcium requirement, the two isoforms showed major differences in their membrane-binding and activation mechanisms. For PKC-alpha, diacylglycerol (DG) specifically enhanced the binding of PKC-alpha to phosphatidylserine (PS)-containing vesicles by 2 orders of magnitude, allowing PKC-alpha high specificity for PS. Also, PKC-alpha could penetrate into the phospholipid monolayer with a packing density comparable to that of the cell membrane only in the presence of Ca2+ and PS. When compared to PKC-alpha, PKC-epsilon had lower binding affinity for PS-containing vesicles both in the presence and in the absence of DG. As a result, PKC-epsilon did not show pronounced specificity for PS. Also, PKC-epsilon showed reduced penetration into PS-containing monolayers, which was comparable to the Ca2+-independent penetration of PKC-alpha into the same monolayers. Taken together, these results suggest the following: (1) The role of Ca2+ in the membrane binding of PKC-alpha is to expose a specific PS-binding site. (2) Once bound to membrane surfaces, PS specifically induces the partial membrane penetration of PKC-alpha that allows its optimal interactions with DG, hence the enhanced membrane binding and activation. (3) PKC-epsilon, due to the lack of Ca2+ binding, cannot specifically interact with PS and DG, which implies the presence of other physiological activator(s) for this isoform.
为阐明传统型和新型蛋白激酶C（PKC）的膜结合及激活机制，我们测定了大鼠PKC-α和PKC-ε与各种组成的磷脂单层及囊泡的相互作用。除了已确定的钙需求差异外，这两种同工型在膜结合和激活机制上也存在主要差异。对于PKC-α，二酰基甘油（DG）特异性地将PKC-α与含磷脂酰丝氨酸（PS）囊泡的结合增强了2个数量级，使PKC-α对PS具有高特异性。此外，只有在Ca2+和PS存在的情况下，PKC-α才能以与细胞膜相当的堆积密度穿透磷脂单层。与PKC-α相比，无论有无DG，PKC-ε对含PS囊泡的结合亲和力都较低。因此，PKC-ε对PS没有明显的特异性。而且，PKC-ε对含PS单层的穿透能力降低，这与PKC-α在相同单层中不依赖Ca2+的穿透能力相当。综上所述，这些结果表明：（1）Ca2+在PKC-α膜结合中的作用是暴露一个特定的PS结合位点。（2）一旦与膜表面结合，PS会特异性地诱导PKC-α部分穿透膜，使其能够与DG实现最佳相互作用，从而增强膜结合和激活。（3）由于缺乏Ca2+结合，PKC-ε不能与PS和DG特异性相互作用，这意味着该同工型存在其他生理激活剂。
