Prior T I, FitzGerald D J, Pastan I
Laboratory of Molecular Biology, DCBDC, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892.
Cell. 1991 Mar 8;64(5):1017-23. doi: 10.1016/0092-8674(91)90325-s.
We have constructed a chimeric toxin composed of Pseudomonas exotoxin A (PE) and the extracellular ribonuclease of Bacillus amyloliquefaciens, barnase. The chimeric protein, termed PE-Bar, reacted with both anti-PE and anti-barnase antisera and had both ADP ribosylation and ribonuclease activities. The chimeric toxin was cytotoxic to the murine fibroblast cell line L929 and to a murine hybridoma resistant to PE. A mutant form of PE-Bar lacking ADP-ribosylating activity was still cytotoxic to L929 cells. Because treatment of cells prelabeld with [3H]uridine resulted in a decrease in their RNA content, we conclude that this cytotoxic effect was due to the ribonuclease activity of barnase molecules that had been translocated to the cytosol. It is now possible to construct chimeric toxins with two or more enzymatic activities that can be delivered to the cytosol of the target cells.
我们构建了一种由铜绿假单胞菌外毒素A（PE）和解淀粉芽孢杆菌的细胞外核糖核酸酶巴那酶组成的嵌合毒素。这种被称为PE - Bar的嵌合蛋白能与抗PE和抗巴那酶抗血清发生反应，并且同时具有ADP核糖基化活性和核糖核酸酶活性。该嵌合毒素对小鼠成纤维细胞系L929以及对PE具有抗性的小鼠杂交瘤具有细胞毒性。一种缺乏ADP核糖基化活性的PE - Bar突变形式对L929细胞仍具有细胞毒性。由于用[³H]尿苷预标记的细胞经处理后其RNA含量降低，我们得出结论，这种细胞毒性作用是由于已转运至胞质溶胶的巴那酶分子的核糖核酸酶活性所致。现在有可能构建具有两种或更多种酶活性的嵌合毒素，这些毒素能够被递送至靶细胞的胞质溶胶中。
