Institute of Microbiology, University of Stuttgart, Allmandring 31, 70569, Stuttgart, Germany.
Institute of Biochemical Engineering, University of Stuttgart, Stuttgart, Germany.
Appl Microbiol Biotechnol. 2020 Nov;104(22):9683-9692. doi: 10.1007/s00253-020-10891-7. Epub 2020 Oct 6.
Agrobacterium tumefaciens synthesizes polyphosphate (polyP) in the form of one or two polyP granules per cell during growth. The A. tumefaciens genome codes for two polyphosphate kinase genes, ppk1 and ppk2, of which only ppk1 is essential for polyP granule formation in vivo. Biochemical characterization of the purified PPK1 and PPK2 proteins revealed a higher substrate specificity of PPK1 (in particular for adenine nucleotides) than for PPK2. In contrast, PPK2 accepted all nucleotides at comparable rates. Most interestingly, PPK2 catalyzed also the formation of tetra-, penta-, hexa-, hepta-, and octa-phosphorylated nucleosides from guanine, cytosine, desoxy-thymidine, and uridine nucleotides and even nona-phosphorylated adenosine. Our data-in combination with in vivo results-suggest that PPK1 is important for the formation of polyP whereas PPK2 has the function to replenish nucleoside triphosphate pools during times of enhanced demand. The potential physiological function(s) of the detected oligophosphorylated nucleotides await clarification. KEY POINTS: •PPK1 and PPK2 have different substrate specificities, •PPK2 is a subgroup 1 member of PPK2s, •PPK2 catalyzes the formation of polyphosphorylated nucleosides.
农杆菌在生长过程中以每个细胞一个或两个多磷酸盐颗粒的形式合成多磷酸盐（polyP）。农杆菌基因组编码两个多磷酸盐激酶基因，ppk1 和 ppk2，其中只有 ppk1 是体内多磷酸盐颗粒形成所必需的。对纯化的 PPK1 和 PPK2 蛋白的生化特性分析表明，PPK1 的底物特异性（特别是对腺嘌呤核苷酸）高于 PPK2。相比之下，PPK2 以可比的速率接受所有核苷酸。最有趣的是，PPK2 还催化鸟嘌呤、胞嘧啶、脱氧胸苷和尿嘧啶核苷酸以及甚至非磷酸化的腺嘌呤核苷酸形成四磷酸、五磷酸、六磷酸、七磷酸和八磷酸化核苷。我们的数据——结合体内结果——表明，PPK1 对于多磷酸盐的形成很重要，而 PPK2 的功能是在需求增加时补充核苷三磷酸池。检测到的低聚磷酸盐核苷酸的潜在生理功能有待阐明。关键点：•PPK1 和 PPK2 具有不同的底物特异性，•PPK2 是 PPK2s 的亚组 1 成员，•PPK2 催化多磷酸化核苷的形成。
