[Cloning and prokaryotic expression of truncated muramidase-released protein gene of human Streptococcus suis type 2].
OBJECTIVE
To clone and express the truncated Habb mrp gene of human Streptococcus suis type 2 (S.suis 2) and detect its activity.


METHODS
A pair of primers based on S.suis 2 mrp gene were schemed out. The turncated mrp (tmrp)gene of S.suis 2 strain Habb isolated from diseased person in Haian, Jiangsu province was cloned and analyzed. The prokaryotic expression plasmid pGEX4T-2-tmrp was constructed.The expression of recombinant protein with glutathione S-transferase (GST) was induced in E.coli TG1. The fusion protein (tMRP-GST) was purified by affinity chromatography, and the GST was cut from tMRP-GST with thrombin protease to gain the truncated MRP (tMRP) antigen. The activity of recombinant protein was analyzed by Western blot.


RESULTS
Sequence analysis showed that the length of the truncated mrp was 957 bp. The prokaryotic expressed production was a fusion protein, whose molecular weight was about 61 kD, and the molecular weight of the purified tMRP protein was about 35 kD. Western blot analysis showed that tMRP-GST and tMRP were detected specifically by MRP antiserum on nitrocellulose membrane.


CONCLUSIONS
The truncated mrp gene of human S.suis 2 strain Habb is successfully cloned, and the high expression of the functional recombinant protein is achieved in the prokaryotic system, which facilitates the further studies on the bio-function and immunology.
Wang
Hai-li
HL
Institute of Military Medical Sciences, Nanjing Command, Nanjing 210002, China. w.haili@163.com
Wang
Chang-jun
CJ
Lu
Cheng-ping
CP
Pan
Xiu-zhen
XZ
Tang
Jia-qi
JQ
D000818:Animals; D015153:Blotting, Western; D003001:Cloning, Molecular; D015870:Gene Expression; D006801:Humans; D009113:Muramidase / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins; D017011:Streptococcus suis / Q000276:immunology / Q000378:metabolism*
D010446:Peptide Fragments; D011994:Recombinant Proteins; D009113:Muramidase