Dankert J R, Esser A F
Department of Comparative and Experimental Pathology, University of Florida, Gainesville 32610.
Biochem J. 1987 Jun 1;244(2):393-9. doi: 10.1042/bj2440393.
The ability of serum complement to kill Gram-negative bacteria requires assembly of the membrane attack complex (MAC) on the cell surface. The molecular events that lead to cell killing after MAC assembly are unknown. We have investigated the effect of C9 on bacterial survival in the presence and absence of its receptor, the C5b-8 complex, on the outer membrane. A fluorescence assay of the membrane potential across the inner bacterial membrane revealed that addition of C9 to cells bearing the performed C5b-8 complex caused a rapid and complete dissipation of the membrane potential. No fluorescence change was observed in serum-resistant strains of Escherichia coli. Addition of trypsin, after C9 was bound to C5b-8, did not rescue the cells from the lethal effects of C9. Furthermore, assays of cell killing kinetics and C9 binding indicate that formation of tubular poly(C9) is not required for killing. When C9 was introduced into the periplasmic space in the absence of its receptor by means of an osmotic shock procedure, cell killing occurred. Other proteins, such as C8 or serum albumin, were not toxic, and C9 was ineffective against two resistant strains. The results presented here and previously [Dankert & Esser (1986) Biochemistry 25, 1094-1100], when considered together, indicate that the 'lethal unit' in complement killing of some Gram-negative bacteria is a C9-derived product that acts by dissipation of cellular energy.
血清补体杀死革兰氏阴性菌的能力需要在细胞表面组装膜攻击复合物（MAC）。MAC组装后导致细胞死亡的分子事件尚不清楚。我们研究了在有或没有其受体（外膜上的C5b-8复合物）存在的情况下，C9对细菌存活的影响。对细菌内膜跨膜电位的荧光测定表明，向带有已形成的C5b-8复合物的细胞中添加C9会导致膜电位迅速且完全消散。在耐血清的大肠杆菌菌株中未观察到荧光变化。在C9与C5b-8结合后添加胰蛋白酶，不能使细胞从C9的致死作用中恢复。此外，细胞杀伤动力学和C9结合测定表明，杀死细菌不需要形成管状多聚（C9）。当通过渗透休克程序在没有其受体的情况下将C9引入周质空间时，会发生细胞杀伤。其他蛋白质，如C8或血清白蛋白，没有毒性，并且C9对两种耐药菌株无效。这里和以前[丹克特和埃塞尔（1986年）《生物化学》25，1094 - 1100]给出的结果综合考虑表明，补体杀死某些革兰氏阴性菌的“致死单位”是一种C9衍生的产物，其作用是消耗细胞能量。
