CLONING AND EXPRESSION OF GENES ENCODING PHEROMONE-INDUCIBLE ANTIGENS OF ENTEROCOCCUS-(STREPTOCOCCUS)-FAECALIS
CLONING AND EXPRESSION OF GENES ENCODING PHEROMONE-INDUCIBLE ANTIGENS OF ENTEROCOCCUS-(STREPTOCOCCUS)-FAECALIS
复制标题
DOI：
10.1128/jb.170.11.5161-5168.1988
复制
发表时间：
1988-11-01
影响因子：
3.2
通讯作者：
DUNNY, GM
中科院分区：
文献类型：
--
作者：
CHRISTIE, PJ;KAO, SM;DUNNY, GM
克隆了58kb碱基粪链球菌四环素耐药质粒pCF10的限制性内切酶片段，并将其导入大肠杆菌和粪肠球菌中，对其编码的信息素诱导接合系统进行了鉴定。免疫印迹分析表明，携带重组质粒pINY1825(克隆的EcoRI C片段)的两个细菌宿主都产生了一个130kDa的抗原，与先前证明参与pCF10介导的信息素诱导的表面排斥的Tra130抗原相同。这两个携带pINY1825的细菌宿主也产生了不同数量的免疫相关118至125 kDa的抗原(指定为Pre-Tra130)，类似于携带pCF10的粪肠球菌细胞产生的抗原。另外一个150 kDa的抗原Tra150可能与信息素诱导的细胞聚集有关，它是由携带pINY1801(克隆的EcoRI C和E片段)的大肠杆菌和粪肠球菌宿主产生的。通过转座子插入突变，将Tra150和Tra130抗原的编码序列进一步定位于pCF10的TRA区。对重组菌株、携带pCF10衍生物的菌株或含有Tn5或Tn917插入片段的各种重组质粒的Western印迹分析表明，包含Tra3至-6片段的pCF10部分(以前由Tn917插入突变定义)包含几个参与调节Tra130和Tra150合成的基因。
Fragments, generated by restriction enzyme digestion, of the 58-kilobase Enterococcus (Streptococcus) faecalis tetracycline resistance plasmid pCF10 were cloned and introduced into Escherichia coli and E. faecalis to characterize the pheromone-inducible conjugation system encoded by this plasmid. Western blot (immunoblot) analyses revealed that a 130-kilodalton (kDa) antigen, identical to the Tra130 antigen shown previously to be involved in pCF10-mediated pheromone-inducible surface exclusion, was produced by both bacterial hosts carrying the recombinant plasmid pINY1825 (cloned EcoRI C fragment). Both bacterial hosts carrying pINY1825 also produced various amounts of immunologically related 118- to 125-kDa antigens (designated pre-Tra130) that resembled antigens produced by E. faecalis cells carrying pCF10. An additional 150-kDa antigen, Tra150, probably involved in pheromone-induced cellular aggregation, was produced by Escherichia coli and E. faecalis hosts carrying pINY1801 (cloned EcoRI C and E fragments). The coding sequences for the Tra150 and Tra130 antigens were further localized in the TRA region of pCF10 by transposon insertion mutagenesis. Western blot analyses of the recombinant strains, and of strains carryng derivatives of pCF10 or various recombinant plasmids containing Tn5 or Tn917 insertions, suggested that the portion of pCF10 comprising the tra3 through -6 segments (previously defined by Tn917 insertions mutagenesis) contained several genes that are involved in regulating the synthesis of Tra130 and Tra150.