沙眼衣原体隐匿质粒编码蛋白的表达及其相互作用

Cryptic plasmid-encoded proteins in Chlamydia trachomatis: expression and interaction

目的分析沙眼衣原体隐匿质粒编码蛋白的相互作用。方法采用基因克隆的方法将沙眼衣原体质粒基因pgp1~4及pgp8分别克隆到pRAC和pRBR上,通过转录激活细菌双杂交系统分析蛋白相互作用。结果 Western blot结果显示Pgp1、Pgp3与大肠杆菌Rpo A氨基末端功能区（α-NTD）融合蛋白及Pgp1、Pgp3、Pgp4与DNA结合蛋白λCI融合蛋白,能在大肠杆菌中表达。在报告菌株中,共表达与α-NTD融合的Pgp3蛋白（α-Pgp3）及与λCI融合的Pgp3蛋白（CI-Pgp3）,导致报告基因产物β-半乳糖苷酶的活性升高。而余无明显变化。结论 pgp3编码的Pgp3能与自身相互作用。

Objective To analyze the interactions among the proteins encoded in the cryptic plasmid of Chlamydia trachomatis （ C. trachomatis）. Methods The DNA fragments of C. trachomatis plasmid genes, pgpl ～ 4 and pgp8, were cloned into the vectors （pRAC and pRBR） , and the target genes were in-frame fused to α subunit N-terminal domain （ot-NTD） and DNA binding protein, CI protein of bacteriophage λ （λCI） , respectively. The expression of cloned genes in Escherichia coli（ E. coli） were determined by Western blotting. The protein-protein interactions were analyzed by a transcription-based bacterial two-hybrid system. Results Western blotting indicated that isopropyl β-D-1-thiogalactopyranoside （IPTG） induced the expression of fusion proteins, α-Pgpl, α-Pgp3, CI-Pgpl, CI-Pgp3 and CI-Pgp4 in E. coli. In the presence of Pgp3 （encoded by plasmid pBRot-Pgp3 ）, the expression of α-Pgp3 and CI-Pgp3 activated the transcription from a reporter gene β-galactosidase in the bacterial two-hybrid assay strain, but Pgp3 had no interaction with other test proteins. Conclusion Pgp3 encoded by pgp3 interacts with Pgp3 in C. trachomatis.