Abstract

Metal ions play an important role in many metabolic processes in all living organisms. At low concentrations, heavy metals such as Fe²⁺, Cu²⁺ and Zn²⁺ are essential cofactors for many enzymes. However, at high concentrations they are toxic. Mesorhizobium species belong to the class α-proteobacteria and have high tolerance to soil acidity, salinity, temperature extremes, and metallicolous conditions. To identify factors responsible for this tolerance we have studied the effects of metal ions on Mesorhizobium delmotii thymidylate kinase (MdTMPK), an essential enzyme in the synthesis of dTTP, thus being vital for cell growth. We show that Mg²⁺ and Mn²⁺ are the divalent metal ions required for catalysis and that Mn²⁺ gives the highest catalytic efficiency. MdTMPK activity in the presence of Mg²⁺ was strongly inhibited by the co-presence of Zn²⁺, Ni²⁺ and Co²⁺. However, the addition of Cs⁺ caused >2-fold enhanced MdTMPK activity. For TMPK from Bacilus anthracis and humans, the effects of Mg²⁺ and Mn²⁺ were similar, whereas the effects of other divalent metal ions were different, and no stimulatory effect of Cs⁺ was observed. Together, our results demonstrate that MdTMPK and BaTMPK function well in the presence of high concentrations of heavy metal ions, introducing a potential contribution of these enzymes to the heavy metal tolerance of Mesorhizobium delmotii and Bacillus anthracis

摘要

金属离子在所有生物体的许多代谢过程中起着重要作用。在低浓度下，Fe ²⁺、Cu ²⁺和 Zn ²⁺等重金属是许多酶必不可少的辅助因子。然而，在高浓度下它们是有毒的。Mesorhizobium物种属于 α-proteobacteria 类，对土壤酸度、盐度、极端温度和金属条件具有高度耐受性。为了确定导致这种耐受性的因素，我们研究了金属离子对德尔莫生根瘤菌胸苷酸激酶 (MdTMPK) 的影响，MdTMPK 是 dTTP 合成中的一种必需酶，因此对细胞生长至关重要。我们表明 Mg ²⁺和 Mn ²⁺是催化所需的二价金属离子，Mn ²⁺提供最高的催化效率。MdTMPK 活性在 Mg ^{2+存在下受到 Zn 2+}、Ni ²⁺和 Co ²⁺共存的强烈抑制。然而，添加 Cs ⁺导致 MdTMPK 活性增强 >2 倍。Mg ²⁺和Mn ^2+对炭疽杆菌和人TMPK的作用相似，而其他二价金属离子的作用不同，Cs ^{+无刺激作用}被观测到。总之，我们的结果表明，MdTMPK 和 BaTMPK 在高浓度重金属离子存在的情况下功能良好，引入了这些酶对Mesorhizobium delmotii 和 Bacillus anthracis的重金属耐受性的潜在贡献