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- Title
结核分枝杆菌转录调控蛋白 Rv0827c(KmtR) 调控靶点的初步研究.
- Authors
王 慧; 崔莹莹; 唐祎依; 吕明月; 党光辉; 崔子寅; 曹 俊; 宋宁宁; 刘思国
- Abstract
Mycobacterium tuberculosis quickly adapts to the host's defense system and environmental changes through the transcriptional regulatory system, resulting in persistent infection, but the mechanism is unclear. In order to study the binding of transcriptional regulatory protein Rv0827c to 23 promoters predicted by the bacteria one-hybrid assay in vivo and in vitro, and to clarify the influencing factors of the binding of Rv0827c to the rv3616c promoter, rv0827c gene was amplified by PCR using MTB H37Ra as a template to construct recombinant plasmid, and the recombinant protein Rv0827c (rRv0827c) was obtained after inducing and purifying. The electrophoretic mobility- shift assay (EMSA) and chromatin immunoprecipitation assay (ChIP) were used to detect the interaction between the rRv0827c protein and the 23 gene promoters and their interaction regions. The results showed that rRv0827c protein can bind to 23 promoters, and it can also specifically bind to the rv3616c (espA) promoter and bind to the major groove region. For metal ions, Ni2+ and Co2+ can inhibit the binding of rRv0827c to rv3616c promoter in vitro, which was confirmed by EMSA, where as Cr3+ plays a positive role. The surface plasmon resonance test (SPR) also further confirmed the binding affinity of rRv0827c and rv3616c promoter. The nucleotide conservation of 23 promoter sequences was analyzed by MEME software, and 4G and 10G were identified as the conserved nucleotide sites for rv3616c to interact with rRv0827c. This study reveals rRv0827c has wide range of potential binding sites and directly binds to the promoter of rv3616c thereby regulating the virulence of Mycobacterium tuberculosis. This provides a new theoretical basis for improving the regulatory network of Rv0827c.
- Subjects
SURFACE plasmon resonance; MYCOBACTERIUM tuberculosis; RECOMBINANT proteins; BINDING sites; PROMOTERS (Genetics)
- Publication
Chinese Journal of Preventive Veterinary Medicine / Zhongguo Yufang Shouyi Xuebao, 2022, Vol 44, Issue 8, p823
- ISSN
1008-0589
- Publication type
Article
- DOI
10.3969/j.issn.1008-0589.202111006