Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI 48824-4320, USA.
Mol Microbiol. 2011 Dec;82(5):1291-300. doi: 10.1111/j.1365-2958.2011.07891.x. Epub 2011 Nov 8.
Nickel is toxic to all forms of life, but the mechanisms of cell damage are unknown. Indeed, environmentally relevant nickel levels (8 µM) inhibit wild-type Escherichia coli growth on glucose minimal medium. The same concentration of nickel also inhibits growth on fructose, but not succinate, lactate or glycerol; these results suggest that fructose-1,6-bisphosphate aldolase (FbaA) is a target of nickel toxicity. Cells stressed by 8 µM Ni(II) for 20 min lost 75% of their FbaA activity, demonstrating that FbaA is inactivated during nickel stress. Furthermore, overexpression of fbaA restored growth of an rcnA mutant in glucose minimal medium supplemented with 4 µM Ni(II), thus confirming that FbaA is a primary target of nickel toxicity. This class II aldolase has an active site zinc and a non-catalytic zinc nearby. Purified FbaA lost 80 % of its activity within 2 min when challenged with 8 µM Ni(II). Nickel-challenged FbaA lost 0.8 zinc and gained 0.8 nickel per inactivated monomer. FbaA mutants (D144A and E174A) affecting the non-catalytic zinc were resistant to nickel inhibition. These results define the primary site of nickel toxicity in E. coli as the class II aldolase FbaA through binding to the non-catalytic zinc site.
镍对所有形式的生命都是有毒的，但细胞损伤的机制尚不清楚。事实上，环境相关的镍水平（8 μM）抑制了野生型大肠杆菌在葡萄糖最小培养基中的生长。同样浓度的镍也抑制了果糖的生长，但不抑制琥珀酸、乳酸或甘油的生长；这些结果表明果糖-1,6-二磷酸醛缩酶（FbaA）是镍毒性的靶标。用 8 μM Ni（II）处理 20 分钟的细胞失去了 75%的 FbaA 活性，表明 FbaA 在镍胁迫下失活。此外，fbaA 的过表达恢复了 rcnA 突变体在葡萄糖最小培养基中的生长，该培养基中补充了 4 μM Ni（II），从而证实 FbaA 是镍毒性的主要靶标。这种 II 类醛缩酶具有一个活性部位锌和一个附近的非催化锌。当用 8 μM Ni（II）挑战时，纯化的 FbaA 在 2 分钟内失去了 80%的活性。受镍挑战的 FbaA 每个失活单体失去 0.8 个锌和获得 0.8 个镍。影响非催化锌的 FbaA 突变体（D144A 和 E174A）对镍抑制具有抗性。这些结果通过结合非催化锌位点，将大肠杆菌中镍毒性的主要靶标定义为 II 类醛缩酶 FbaA。
