Mold C, Walter E I, Medof M E
Department of Microbiology, University of New Mexico, Albuquerque 87131.
J Immunol. 1990 Dec 1;145(11):3836-41.
Decay-accelerating factor (DAF) is a C regulatory protein which functions in membranes to inhibit autologous C activation on cell surfaces. A liposome model was used to study the mechanism of DAF action and examine the effects of membrane-bound glycophorin and LPS on the regulatory activity of DAF. Liposomes were incubated in MgEGTA-treated human serum and activation of the alternative pathway measured by C3b binding. Liposomes composed of phosphatidylcholine, phosphatidylethanolamine, and cholesterol activated the alternative pathway in proportion to their content of PE. Incorporation of 10(-7) mol/mol phospholipid of either human E or HeLa cell-derived DAF inhibited C activation by liposomes containing 40% phosphatidylethanolamine by 50%, an efficiency comparable to that observed in intact E. HeLa DAF that had been treated with phosphatidylinositol-specific phospholipase C to remove its glycolipid anchor had no effect on C activation by liposomes at concentrations as high as 10(-5) mol/mol phospholipid. Incorporation of DAF into liposomes prepared with bound C3b inhibited the deposition of additional C3b by C3bBbP. However, the incorporated DAF increased the amount of Bb generated from B in the presence of D indicating that accelerated decay of the convertase was the primary effect of DAF. Similarly, treatment of intact human E with anti-DAF decreased the amount of Bb generated by the alternative pathway convertase. To study the effects of other membrane components on DAF activity, liposomes were prepared with purified human glycophorin A or LPS. In glycophorin liposomes the presence of PE was required to activate the alternative pathway and DAF inhibited this activation. In contrast, LPS liposomes bound C3b independently of PE and the incorporation of DAF had no effect. These results demonstrate that within a membrane, DAF's inhibitory activity on the alternative pathway C3 convertase is mediated independently of other membrane proteins, that in this model the major activity of DAF is to accelerate convertase decay, and that the presence of other membrane molecules that may serve as C3 acceptors can circumvent DAF function.
衰变加速因子（DAF）是一种C调节蛋白，在细胞膜中发挥作用，抑制细胞表面自身补体的激活。采用脂质体模型研究DAF的作用机制，并检测膜结合血型糖蛋白和脂多糖对DAF调节活性的影响。将脂质体置于经MgEGTA处理的人血清中孵育，通过C3b结合检测替代途径的激活情况。由磷脂酰胆碱、磷脂酰乙醇胺和胆固醇组成的脂质体，其替代途径的激活程度与其PE含量成正比。每摩尔磷脂中掺入10^(-7)摩尔人E或HeLa细胞来源的DAF，可使含40%磷脂酰乙醇胺的脂质体的C激活受到50%的抑制，其效率与在完整E细胞中观察到的相当。用磷脂酰肌醇特异性磷脂酶C处理以去除其糖脂锚定的HeLa DAF，在高达每摩尔磷脂10^(-5)摩尔的浓度下，对脂质体的C激活没有影响。将DAF掺入用结合的C3b制备的脂质体中，可抑制C3bBbP沉积额外的C3b。然而，掺入的DAF在有D存在的情况下增加了由B产生的Bb的量，表明转化酶的加速衰变是DAF的主要作用。同样，用抗DAF处理完整的人E细胞，可减少替代途径转化酶产生的Bb的量。为了研究其他膜成分对DAF活性的影响，用纯化的人血型糖蛋白A或脂多糖制备脂质体。在血型糖蛋白脂质体中，需要有PE才能激活替代途径，而DAF可抑制这种激活。相反，脂多糖脂质体独立于PE结合C3b，掺入DAF没有影响。这些结果表明，在细胞膜内，DAF对替代途径C3转化酶的抑制活性独立于其他膜蛋白介导，在该模型中DAF的主要活性是加速转化酶衰变，并且其他可能作为C3受体的膜分子的存在可以规避DAF的功能。
