[Expression and identification of gene Loa22 from the virulent strain of serovar Lai of L. interrogans and its cytotoxicity to macrophages].
OBJECTIVE
To study the function of Loa22 gene from virulent serovar Lai. L. interrogans by expressing its protein.


METHODS
The recombinant plasmid with Loa22 was constructed and its expression was induced by IPTG. The expression product was identified by SDS-PAGE and Western Blotting and purified with GST-affinity chromatography. The purified protein was applied to mice macrophages ANA-1 cells purified by GST-affinity column and exerted to ANA-1 cells. The cytotoxicity of purified protein to ANA-1 was evaluated by detecting LDH activity in culture medium, XTT aborbtion and apoptosis ration.


RESULTS
The recombinant plasmid with Loa22 mature peptide was constructed successfully and the protein was purified subsequently. Significant higher level of LDH activity and apoptosis ratio and lower XTT absorption were observed by comparison of expression protein treated cells and those untreated.


CONCLUSIONS
Loa22 had a toxic effect on ANA-1 and the gene Loa22 might be a virulent-related gene of L. interrogans.
Research Unit of Infection and Immunity, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Ying
Y
Zhang
Lang
L
Bao
Hai-Long
HL
Zhu
Bi
B
Huang
Le
L
Zhang
Hui-Dong
HD
Zhang
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology; D015153:Blotting, Western; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D007919:Leptospira / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D010957:Plasmids / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000494:pharmacology; D014774:Virulence
D001425:Bacterial Outer Membrane Proteins; D011994:Recombinant Proteins; C078167:OMPA outer membrane proteins; D007770:L-Lactate Dehydrogenase