Lukac M, Pier G B, Collier R J
Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, Massachusetts.
Infect Immun. 1988 Dec;56(12):3095-8. doi: 10.1128/iai.56.12.3095-3098.1988.
Glutamic acid-553 of Pseudomonas aeruginosa exotoxin A (ETA), identified previously as an active-site residue, was deleted by oligonucleotide-directed mutagenesis of the cloned toxin gene in Escherichia coli. The purified mutant toxin was stable, fully immunoreactive, and capable of blocking toxin receptors. ADP-ribosyltransferase and cytotoxic activities were at least 10(6)-fold lower than those of wild-type ETA, and injection of mice with 50 micrograms (equivalent to 400 lethal doses of ETA) produced no ill effects. The mutant toxin elicited high levels of neutralizing anti-ETA antibodies in mice, which protected against a challenge with 100 micrograms of authentic ETA (greater than 600 lethal doses). The mutant protein has the attributes of a toxoid and may be useful as a component of vaccines for individuals at risk for infection by P. aeruginosa.
铜绿假单胞菌外毒素A（ETA）的谷氨酸-553先前被鉴定为活性位点残基，通过对大肠杆菌中克隆的毒素基因进行寡核苷酸定向诱变将其删除。纯化的突变毒素稳定，具有完全免疫反应性，并且能够阻断毒素受体。ADP-核糖基转移酶和细胞毒性活性比野生型ETA至少低10^6倍，给小鼠注射50微克（相当于400个ETA致死剂量）未产生不良影响。突变毒素在小鼠体内引发了高水平的中和抗ETA抗体，可保护小鼠免受100微克天然ETA（大于600个致死剂量）的攻击。该突变蛋白具有类毒素的特性，可能作为疫苗成分用于有感染铜绿假单胞菌风险的个体。
