Rebecchi M, Peterson A, McLaughlin S
Department of Physiology and Biophysics, State University of New York, Stony Brook 11794-8661.
Biochemistry. 1992 Dec 29;31(51):12742-7. doi: 10.1021/bi00166a005.
We studied the binding of phosphoinositide-specific phospholipase C-delta 1 (PLC-delta) to vesicles containing the negatively charged phospholipids phosphatidylinositol 4,5-bisphosphate (PIP2) and phosphatidylserine (PS). PLC-delta did not bind significantly to large unilamellar vesicles formed from the zwitterionic lipid phosphatidylcholine (PC) but bound strongly to vesicles formed from mixtures of PC and PIP2. The apparent association constant for the putative 1:1 complex formed between PLC-delta and PIP2 was Ka congruent to 10(5) M-1. The binding strength increased further (Ka congruent to 10(6) M-1) when the vesicles also contained 30% PS. High-affinity binding of PLC-delta to PIP2 did not require Ca2+. PLC-delta bound only weakly to vesicles formed from mixtures of PC and either PS or phosphatidylinositol (PI); binding increased as the mole fraction of acidic lipid in the vesicles increased. We also studied the membrane binding of a small basic peptide that corresponds to a conserved region of PLC. Like PLC-delta, the peptide bound weakly to vesicles containing monovalent negatively charged lipids; unlike PLC-delta, it did not bind strongly to vesicles containing PIP2. Our data suggest that a significant fraction of the PLC-delta in a cell could be bound to PIP2 on the cytoplasmic surface of the plasma membrane.
我们研究了磷酸肌醇特异性磷脂酶C-δ1（PLC-δ）与含有带负电荷磷脂磷脂酰肌醇4,5-二磷酸（PIP2）和磷脂酰丝氨酸（PS）的囊泡的结合情况。PLC-δ与由两性离子脂质磷脂酰胆碱（PC）形成的大单层囊泡没有明显结合，但与由PC和PIP2混合物形成的囊泡有强烈结合。PLC-δ与PIP2形成的假定1:1复合物的表观缔合常数Ka约为10⁵ M⁻¹。当囊泡中还含有30%的PS时，结合强度进一步增加（Ka约为10⁶ M⁻¹）。PLC-δ与PIP2的高亲和力结合不需要Ca²⁺。PLC-δ与由PC和PS或磷脂酰肌醇（PI）混合物形成的囊泡仅有微弱结合；随着囊泡中酸性脂质摩尔分数的增加，结合增强。我们还研究了与PLC保守区域对应的小碱性肽的膜结合情况。与PLC-δ一样，该肽与含有单价负电荷脂质的囊泡结合较弱；与PLC-δ不同的是，它与含有PIP2的囊泡没有强烈结合。我们的数据表明，细胞中相当一部分PLC-δ可能与质膜细胞质表面的PIP2结合。
