Immunological characteristics of the PhoP protein of two-component system in Mycobacterium tuberculosis
10.3969/j.issn.1002-2694.2024.00.051
- VernacularTitle:结核分枝杆菌双组分系统PhoP蛋白的免疫学特性
- Author:
Xue LI
1
,
2
;
Huan-Huan NING
;
Jian KANG
;
Ming-Ze XU
;
Ruo-Nan CUI
;
Ting DAI
;
Yan-Zhi LU
;
Sa XUE
;
Yin-Lan BAI
Author Information
1. 西北大学生命科学学院,西安 710069
2. 空军军医大学基础医学院微生物与病原生物学教研室,西安 710032
- Keywords:
Mycobacterium tuberculosis;
two-component system;
PhoP protein;
humoral immune response;
cellular immune response
- From:
Chinese Journal of Zoonoses
2024;40(4):352-358
- CountryChina
- Language:Chinese
-
Abstract:
In this study,the immunological characteristics of the PhoP protein were explored with a two-component system of Mycobacterium tuberculosis(Mtb).Bioinformatics was used to predict the B and T cell epitopes of the PhoP protein.A re-combinant expression plasmid was constructed by PCR analysis of the phoP sequence and cloning into the prokaryotic expres-sion vector pET-28a(+).Competent Escherichia coli BL21 cells were transformed with the recombinant plasmid and expres-sion was induced with IPTG.The recombinant PhoP protein was purified by affinity chromatography.Serum levels of PhoP-specific antibodies in Mtb-infected mice and tuberculosis(TB)patients were analyzed with an ELISA.BALB/c mice were im-munized with the PhoP recombinant protein by intramuscular injection.Sera of mice were collected and antibody titers were detected with an ELISA and specificity was assessed by West-ern blot analysis.Mouse splenocytes were isolated and the pro-portions of IFN-y-positive cells and cytokine levels were detec-ted with an ELISpot and ELISA,respectively.Bioinformatics i-dentified 24 B cell and 11 T cell epitopes of the PhoP protein.A prokaryotic recombinant vector of PhoP was successfully con-structed and the recombinant PhoP protein was obtained by purification.Specific antibody levels to PhoP in sera of Mtb in-fected mice and TB patients increased significantly,with preci-sion of 99.9％and 82.5％,and specificity of 100％,respectively.PhoP protein immunization successfully induced production of specific antibodies in mice.Stimulated by antigens in vitro,IL-2 and IFN-γ levels were significantly increased in the splenocytes of immunized mice.Immunization with the PhoP protein induce a humoral immune response and Thl-dominated cellular immu-nity,indicating that the PhoP protein was immunogenic with diagnostic efficacy for TB.These results lay a foundation to clari-fy the role of PhoP in Mtb infection and application for diagnosis and prevention of TB.
