Cobra venom cytotoxin free of phospholipase a, and pts effect on model membranes and T leukemia cells
Cobra venom cytotoxin free of phospholipase a, and pts effect on model membranes and T leukemia cells
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DOI：
10.1007/s002329900165
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发表时间：
1997-01-15
影响因子：
2.4
通讯作者：
Rael, ED
中科院分区：
文献类型：
--
作者：
Gasanov, SE;Alsarraj, MA;Rael, ED
蛇毒的膜活性毒素以前曾被用于研究蛋白质-脂质相互作用和探测生物膜的物理和生化状态。为了扩展这些研究，我们从眼镜蛇毒中分离出一种不含磷脂酶A（2）（PLA（2））的细胞毒素。的氨基酸组成，pI（10.2），和净电荷的细胞毒素相比，以及与其他眼镜蛇毒液分离的膜活性毒素。自旋标记法显示，在pH值为7.0 - 5.0的缓冲液中，细胞毒素与PLA（2）缔合，但在pH 4.0时不缔合。结果表明，在天然蛇毒中，细胞毒素与PLA（2）（pI接近4.8）静电缔合。用自旋探针在取向脂质多层膜中的EPR和超声处理脂质体的H-1-NMR研究了细胞毒素对模型磷脂膜的影响。细胞毒素没有显着影响包装的脂质在纯磷脂酰胆碱（PC）膜和PC膜含有10摩尔%的磷脂酸（PA）或心磷脂（CL）。然而，细胞毒素诱导的PC膜含有40摩尔%的PA或CL的膜通透性和非双层结构的形成增加。纯化的细胞毒素对Jurkat细胞具有杀细胞作用，但对正常人淋巴细胞的作用不大。然而，当用10（-9）M PLA（2）和10（-5）M细胞毒素联合处理时，Jurkat细胞和正常淋巴细胞被等同地杀死。从其对模型膜和Jurkat细胞的影响，它建议，纯化的细胞毒素优先目标和破坏膜，是富含酸性磷脂的细胞外侧的质膜。
Membrane-active toxins from snake venom have been used previously to study protein-lipid interactions and to probe the physical and biochemical states of biomembranes. To extend these studies, we have isolated from Naja naja kaowthia (cobra) venom a cytotoxin free of detectable phospholipase A(2) (PLA(2)). The amino acid composition, pI (10.2), and net charge of the cytotoxin compares well with membrane-active toxins isolated from venoms of other cobras. The cytotoxin, shown by a spin label method, associates with PLA(2) in buffers at pH values between 7.0 and 5.0, but not at pH 4.0. It is suggested that cytotoxin and PLA(2) (pI close to 4.8) associate electrostatically in the native venom. The effect of the cytotoxin on model phospholipid membranes was studied by EPR of spin probes in oriented lipid multilayers and H-1-NMR of sonicated liposomes. The cytotoxin did not significantly affect the packing of lipids in pure phosphatidylcholine (PC) membranes and in PC membranes containing 10 mol% phosphatidic acid (PA) or cardiolipin (CL). However, the cytotoxin induced an increase in membrane permeability and formation of nonbilayer structures in PC membranes containing 40 mol% of PA or CL. The purified cytotoxin was cytocidal to Jurkat cells, but had little effect on normal human lymphocytes. However, both Jurkat cells and normal lymphocytes were killed equivalently when treated with 10(-9) M PLA(2) and 10(-5) M cytotoxin in combination. From its effect on model membranes and Jurkat cells, it is suggested that purified cytotoxin preferentially targets and disrupts membranes that are rich in acidic phospholipids on the extracellular side of the plasma membrane.