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Updated: Jun 9, 2026
Mycobacterium tuberculosis Extracellular Vesicle Enrichment through Size Exclusion Chromatography
Published on: May 19, 2022
[Expression, purification, and characterization Mycobacterium tuberculosis Rv1168c]
Xiaoli Yu1, Zhanqiang Sun, Chenjun Zhou
1School of Biolgy and Pharmaceutical Engineering, Wuhan Polytechnic University, Wuhan 430023, China. yxll268@126.com
Objective:
To express and purify the Pro-Pro-Glu (PPE) family protein Rv1168c of Mycobacterium tuberculosis in E. coli. and to study the structure of Rv1168c.
Methods:
The Rv1168c gene was amplified by PCR from Mycobacterium tuberculosis H37Rv strain genomic DNA and cloned into a prokaryotic expression vector pET32a The resulting recombinant expression plasmid pET32a-Rv1168c was then transformed into the E. coli strain DH5alpha and a high-level expression E. coli BL21(DE3) was established after induction with Isopropyl-beta-D-thiogalactopyranoside (IPTG). SDS-PAGE and mass spectrum analysis determined the relative molecular weight of this recombinant Rv1168c protein. It's secondary and 3D structures were determined by circular dichroism and homologous modeling.
Results:
The Mycobacterium tuberculosis Rv1168c gene (971bp) and high purified recombinant Rv1168c protein was obtained. The relative molecular weight of recombinant Rv1168c protein was determined to be 51.5 kDa (vector included). Secondary structure of Rv1168c had about 34.4% alpha helix, 33.7%, beta tune, 31.9% random coil at 25 delta C. Homologous modeling shows Rv1168c as (beta/alpha)5 protein.
Conclusion:
This study obtained purified recombinant Rv1168c protein and laid the foundation for exploration of the relationship between the structure and function of Rv1168c in the tuberculosis.
Insights
Researchers successfully expressed and purified the Pro-Pro-Glu (PPE) family protein Rv1168c from Mycobacterium tuberculosis in E. coli. This study provides purified recombinant Rv1168c protein for further functional and structural analysis in tuberculosis research.
Area of Science:
- Molecular Biology
- Structural Biology
- Microbiology
Background:
- The Pro-Pro-Glu (PPE) family protein Rv1168c is a component of Mycobacterium tuberculosis.
- Understanding the structure and function of Rv1168c is crucial for tuberculosis research.
Purpose of the Study:
- To express and purify the Rv1168c protein from Mycobacterium tuberculosis in E. coli.
- To investigate the secondary and three-dimensional structure of Rv1168c.
Main Methods:
- Amplification and cloning of the Rv1168c gene into a pET32a prokaryotic expression vector.
- High-level expression of recombinant Rv1168c in E. coli BL21(DE3) induced by Isopropyl-beta-D-thiogalactopyranoside (IPTG).
- Analysis of protein molecular weight by SDS-PAGE and mass spectrometry.
- Determination of secondary and 3D structures using circular dichroism and homologous modeling.
Main Results:
- Successfully obtained the Mycobacterium tuberculosis Rv1168c gene (971bp) and highly purified recombinant Rv1168c protein.
- The relative molecular weight of the recombinant Rv1168c protein (including the vector) was determined to be 51.5 kDa.
- Secondary structure analysis revealed approximately 34.4% alpha helix, 33.7% beta-sheet, and 31.9% random coil at 25°C.
- Homologous modeling predicted Rv1168c to be a (beta/alpha)5 protein.
Conclusions:
- Purified recombinant Rv1168c protein was successfully obtained.
- This study provides a foundation for further investigation into the structure-function relationship of Rv1168c in the context of tuberculosis.
