[Expression, purification, and characterization Mycobacterium tuberculosis Rv1168c].
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.


CONCLUSIONS
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.
Yu
Xiaoli
X
School of Biolgy and Pharmaceutical Engineering, Wuhan Polytechnic University, Wuhan 430023, China. yxll268@126.com
Sun
Zhanqiang
Z
Zhou
Chenjun
C
Wen
Zilu
Z
Chen
Jun
J
Sun
Qingwen
Q
Wang
Honghai
H
Zhang
Shulin
S
D000942:Antigens, Bacterial / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D008970:Molecular Weight; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000378:metabolism*; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
D000942:Antigens, Bacterial; D011994:Recombinant Proteins; C530985:Rv1168c protein, Mycobacterium tuberculosis