Characterization of Agrobacterium tumefaciens PPKs reveals the formation of oligophosphorylated products up to nucleosid
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BIOTECHNOLOGICALLY RELEVANT ENZYMES AND PROTEINS
Characterization of Agrobacterium tumefaciens PPKs reveals the formation of oligophosphorylated products up to nucleoside nona-phosphates Celina Frank 1 & Attila Teleki 2 & Dieter Jendrossek 1 Received: 13 July 2020 / Revised: 28 August 2020 / Accepted: 4 September 2020 # The Author(s) 2020
Abstract 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, ppk1AT and ppk2AT, of which only ppk1AT is essential for polyP granule formation in vivo. Biochemical characterization of the purified PPK1AT and PPK2AT proteins revealed a higher substrate specificity of PPK1AT (in particular for adenine nucleotides) than for PPK2AT. In contrast, PPK2AT accepted all nucleotides at comparable rates. Most interestingly, PPK2AT catalyzed also the formation of tetra-, penta-, hexa-, hepta-, and octa-phosphorylated nucleosides from guanine, cytosine, desoxy-thymidine, and uridine nucleotides and even nonaphosphorylated adenosine. Our data—in combination with in vivo results—suggest that PPK1AT is important for the formation of polyP whereas PPK2AT 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 •PPK1AT and PPK2AT have different substrate specificities, •PPK2AT is a subgroup 1 member of PPK2s, •PPK2AT catalyzes the formation of polyphosphorylated nucleosides Keywords Polyphosphate . Polyphosphate kinase . Agrobacterium tumefaciens . Nucleotides
Introduction Polyphosphate (polyP) is an inorganic polymer in which phosphate residues are linked by energy-rich phosphoanhydride bonds. It can be formed either abiotically (vulcanism) or biologically by the action of polyP kinases (PPKs in prokaryotes) or other enzymes (in eukaryotes). PolyP is ubiquitously distributed in species of all domains of Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00253-020-10891-7) contains supplementary material, which is available to authorized users. * Dieter Jendrossek [email protected] 1
Institute of Microbiology, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany
2
Institute of Biochemical Engineering, University of Stuttgart, Stuttgart, Germany
life (Kornberg et al. 1999; Rao et al. 2009; Kulakovskaya et al. 2014). In most prokaryotes, polyP is present in form of granule-like inclusions (polyP granules or voluntin granules) with diameters mostly in the range of 50 to 200 nm. In yeasts, polyP can be accumulated up to ≈ one quarter of the cellular weight in vacuole-like compartments (Christ et al. 2020a). PolyP represents a reservoir of phosphorous and apparently is also involved in various forms of stress resistance such as tolerance against heavy metals, elevated temperature, or reactive oxygen species and can fulfill functi
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