Mosior M, Epand R M
Department of Biochemistry, McMaster University, Hamilton, Ontario, Canada.
J Biol Chem. 1994 May 13;269(19):13798-805.
Association of calcium-dependent isotypes of protein kinase C (PKC) with a phospholipid bilayer is regulated by a single Ca(2+)-binding site. The dependence of PKC association with phosphatidylserine-containing membranes on the concentration of Ca2+ is linear in the submicro- to submillimolar range. The Ca(2+)-regulated association of PKC with the membrane is sensitive to the factors that alter the diffuse double-layer potential produced by anionic lipids such as phosphatidylserine (PS). This indicates that the Ca(2+)-binding site on the membrane-bound enzyme senses a higher concentration of Ca2+ than is present in bulk solution. This is a consequence of the accumulation of Ca2+ in the layer adjacent to the plane of the membrane by the double-layer potential. Calculations based on the Gouy-Chapman-Stern theory of the diffuse double layer yielded a unique value of the Ca2+ dissociation constant for the Ca(2+)-PKC-bilayer complex equal to approximately 700 nM. The soluble form of the enzyme has a 3.5 order of magnitude lower affinity for Ca2+. The free energy of interaction between the Ca(2+)- and PS-binding sites is large (approximately 5 kcal/mol). In contrast, the interaction between the diacylglycerol-binding site and either the Ca(2+)- or PS-binding site appears to be weak.
蛋白激酶C（PKC）的钙依赖性同工型与磷脂双层的结合受单个Ca(2+)结合位点调控。在亚微摩尔至亚毫摩尔范围内，PKC与含磷脂酰丝氨酸的膜的结合对Ca2+浓度的依赖性呈线性。PKC与膜的Ca(2+)调节结合对改变由磷脂酰丝氨酸（PS）等阴离子脂质产生的扩散双层电位的因素敏感。这表明膜结合酶上的Ca(2+)结合位点感知到的Ca2+浓度高于本体溶液中的浓度。这是双层电位使Ca2+在膜平面相邻层中积累的结果。基于扩散双层的 Gouy-Chapman-Stern理论的计算得出Ca(2+)-PKC-双层复合物的Ca2+解离常数的唯一值约为700 nM。该酶的可溶性形式对Ca2+的亲和力低3.5个数量级。Ca(2+)和PS结合位点之间的相互作用自由能很大（约5千卡/摩尔）。相比之下，二酰基甘油结合位点与Ca(2+)或PS结合位点之间的相互作用似乎较弱。
