Reversion of recombinant toxoids: mutations in diphtheria toxin that partially compensate for active-site deletions.
Reversion of recombinant toxoids: mutations in diphtheria toxin that partially compensate for active-site deletions.
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重组类毒素的逆转：白喉毒素的突变部分补偿了活性位点的缺失。
DOI：
10.1073/pnas.89.13.6207
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发表时间：
1992
影响因子：
11.1
通讯作者：
Collier,RJ
中科院分区：
文献类型：
--
作者：
Killeen,KP;Escuyer,V;Mekalanos,JJ;Collier,RJ
删除白喉毒素的一个重要活性位点残基谷氨酸-148，会将该毒素的 ADP-核糖基转移酶活性降低 10(4) 倍以上。我们考虑使用这种突变来构建重组类毒素，用于减毒活疫苗的表达，并探索可能导致回复的第二位点突变。通过用谷氨酸取代缬氨酸 147 或通过向 NH2 末端延伸五个残基的缺失，从而使谷氨酸 142 紧邻酪氨酸 149，活性得到部分恢复。在这两种突变体中，所示的谷氨酸可能占据与未突变蛋白中谷氨酸148相似的空间位点。因此，简单地删除一个关键残基并不能保证第二位点突变不能轻易恢复类毒素的活性。
Deleting an important active-site residue of diphtheria toxin, glutamic acid-148, reduces the toxin's ADP-ribosyltransferase activity by a factor of greater than 10(4). We considered using this mutation to construct a recombinant toxoid for expression by live attenuated vaccines and explored second-site mutations that might cause reversion. Activity was partially restored by substituting glutamic acid for valine-147 or by extending the deletion by five residues toward the NH2 terminus, thereby placing glutamic acid-142 immediately adjacent to tyrosine-149. In both mutants the indicated glutamic acid may occupy a spatial locus similar to that of glutamic acid-148 in the unmutated protein. Simply deleting a crucial residue does not, therefore, provide confidence that a second-site mutation could not readily restore activity to a toxoid.