甜菜BvMTP11基因在大肠杆菌中的镉耐受功能分析

Functional analysis of Beta vulgaris BvMTP11 gene in E. coli under cadmium stress

为了研究甜菜BvMTP11基因(LOC104886952)在重金属Cd逆境胁迫过程中的分子功能,明确其在细胞重金属解毒中的作用,本研究将BvMTP11基因构建于E.coli原核表达载体pEASY-Blunt E1上,并将该重组质粒转化到大肠杆菌BL21中。SDS-PAGE电泳表明,通过1 mmol·L-1 IPTG的诱导,在大肠杆菌总蛋白46 kDa左右的位置上,获得了重组菌表达的his tag-BvMTP11的融合蛋白。当施加0.5 mmol·L-1的CdCl2重金属逆境胁迫后,对照菌的生长受到了较为严重的抑制。相反,过量表达BvMTP11蛋白的重组菌表现出了较好的生长状态,并且其体内的镉离子含量也要远低于对照菌。这说明,甜菜BvMTP11基因可以在一定程度上提高E.coli对Cd2+的耐受性,并有可能通过对Cd2+的转运和富集来解毒重金属对细胞的伤害。研究结果也进一步说明了甜菜BvMTP11在镉逆境胁迫分子机制中发挥着重要作用。

In order to investigate the molecular function of sugar beet BvMTP11 gene(LOC104886952) in the process of cadmium stress, and clarify its role in the detoxification of heavy metals in cells, this study constructed BvMTP11 gene into E. coli prokaryotic expression vector pEASY-Blunt E1, and the recombinant plasmid was transformed into E. coli BL21. SDS-PAGE showed that, after 1 mmol·L-1IPTG induction, the fusion protein of his tag-BvMTP11 was obtained by a band of 46 kDa of total E. coli protein.Under 0.5 mmol·L-1 CdCl2 heavy metal stress, the growth of the control cells was inhibited seriously;on the contrary, the recombinant bacteria over-expressing BvMTP11 exhibited a better growth state, and its concentration of Cd2+ was much lower than that of the control bacteria. The results indicated that sugar beet BvMTP11 gene could improve cadmium tolerance of E. coli to some extent, and might detoxify the damage of heavy metal to cells through transportation and extracellular excretion of Cd2+. The results also suggested that BvMTP11 played an essential role in the molecular mechanism of cadmium stress.