Marchitto K S, Munoz J J, Keith J M
Infect Immun. 1987 May;55(5):1309-13. doi: 10.1128/iai.55.5.1309-1313.1987.
Monoclonal antibodies with specificity for pertussis toxin subunits S1, S2, and S4 were used in Western blots to show that the subunits were not secreted into culture medium from Tn5 insertion mutants. The mutants are deficient in toxin biological activities due to an insertion in the S3 subunit structural gene. The Western blots demonstrated that each of the respective subunits was exported in a wild-type strain. Anti-S1 and anti-S2 monoclonal antibodies were capable of detecting subunits in solubilized whole-cell material from a wild-type strain and from the Tn5 mutants lacking only in biologically active toxin (Tox-). Another Tn5 insertion mutant, lacking all known B. pertussis virulence factors (Vir-), did not produce any of the subunits either in whole cellular extracts or in culture supernatants. The data demonstrate that Tn5 Tox- insertion mutants, though defective in toxin activity, synthesize some toxin subunits. The presence of the S3 subunit is most likely a necessity for transport of the toxin from cells. Alternatively, a nonstructural gene coding for a protein involved in transport of the toxin across the membrane may be affected by the Tn5 mutation.
用对百日咳毒素亚基S1、S2和S4具有特异性的单克隆抗体进行蛋白质印迹分析，以表明这些亚基不会从Tn5插入突变体分泌到培养基中。由于S3亚基结构基因中的插入，这些突变体缺乏毒素生物学活性。蛋白质印迹分析表明，野生型菌株中各自的亚基均被输出。抗S1和抗S2单克隆抗体能够检测来自野生型菌株和仅缺乏生物活性毒素（Tox-）的Tn5突变体的溶解全细胞材料中的亚基。另一个Tn5插入突变体，缺乏所有已知的百日咳博德特氏菌毒力因子（Vir-），在全细胞提取物或培养上清液中均不产生任何亚基。数据表明，Tn5 Tox-插入突变体虽然毒素活性有缺陷，但能合成一些毒素亚基。S3亚基的存在很可能是毒素从细胞中转运的必要条件。或者，编码参与毒素跨膜转运的蛋白质的非结构基因可能受到Tn5突变的影响。
