Expression and purification of a jasmonic acid carboxyl methyltransferase from Salvia miltiorrhiza in E.coli
Yao, N.; Zheng, H.; Jing, L.; Ma, L.-g.; Shen, Y.; Chen, M.
Yao Xue Xue Bao 51(10): 1643-1650
2016
ISSN/ISBN: 0513-4870 PMID: 29932621 Document Number: 688571
Jasmonic acid carboxyl methyltransferase（JMT）, a key enzyme for jasmonate（JA） biosynthesis, catalyzes the methylation of JA to form Me JA. To characterize the function of JMT, a plasmid pGEX-4T- SmJMT1 harboring JMT1（SmJMT1） gene from Salvia miltiorrhiza was successfully transformed into E. coli BL21（DE3） for protein expression. The recombination SmJMT1 was separated using SDS-PAGE and the size of expressed SmJMT1 protein was consistent with the prediction. The bacterial growth conditions were determined for optimal expression, which include growth temperature, incubation time, IPTG concentrations and culture density. The optimal growth conditions for SmJMT1 were that the bacterial cultures were grown to an A600 of 0.8, and induced with IPTG at a final concentration of 0.4 mmol·L-1, and then incubated for 8 h at 20 ℃. The expression of SmJMT1 in E. coli was confirmed by Western blotting, and mass spectrometry analysis of methyltransferase family. The successful expression and purification of JMT in this study provide the basis for more study of JA biosynthetic pathway and JA-regulated secondary metabolism of medicinal plants.