[Cloning and expression of the nickel/cobalt transferase gene in E. coli BL21 and bioaccumulation of nickel ion by genetically engineered strain].
1 053 bp of the nickel/cobalt transferase gene, NiCoT gene, from Staphylococcus aureus ATCC6538 was amplified by PCR and ligated into vector pET-3c. The recombined plasmid was constructed and transferred into E. coli BL21 at appropriate temperature. The recombined strain was isolated and identified by restriction enzyme digestion and PCR amplification. Nucleotide sequence analysis showed that the identity was more than 97% between the nickel/cobalt transferase gene from S. aureus ATCC6538 and the reported gene sequence of other species or subspecies of S. aureus at GenBank. There was a characteristic protein strip near the relatively molecular weight of 39 000 in the SDS-PAGE picture, which was identical to the expected value. The result demonstrated that the NiCoT gene of S. aureus had been successfully expressed in E. coli BL21. The E. coli BL21 containing the NiCoT gene had the highest bioaccumulation quantity when induced with 1.00 mmol x L(-1) IPTG for 4 h. The quantity of equilibrium accumulation of the genetically engineered E. coli BL21 was 11.33 mg x g(-1), which was 3 times more than that of the original E. coli BL21 at different nickel concentrations. The NiCoT of S. aureus ATCC6538 was a highly selective and accumulative nickel transporter and belonged to the class III of the nickel/cobalt transferase.
Zhang
Ying-Ming
YM
Department of Environmental Engineering, Jinan University, Guangzhou 510632, China. zym-333@sohu.com
Yin
Hua
H
Ye
Jin-Shao
JS
Peng
Hui
H
Zhang
Na
N
Qin
Hua-Ming
HM
Yang
Feng
F
He
Bao-Yan
BY
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D003035:Cobalt / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D005818:Genetic Engineering; D009532:Nickel / Q000378:metabolism*; D013211:Staphylococcus aureus / Q000201:enzymology* / Q000235:genetics; D014166:Transferases / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D003035:Cobalt; D009532:Nickel; D014166:Transferases