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  • Vidilaseris, Keni; Kellosalo, Juho; Goldman, Adrian (2018)
    Membrane-bound pyrophosphatases (mPPases) are homodimeric integral membrane proteins that hydrolyse pyrophosphate into orthophosphates coupled to the active transport of protons or sodium ions across membranes. They occur in bacteria, archaea, plants, and protist parasites. As they are essential in protist parasites and there are no homologous proteins in animals and humans, these enzymes represent an excellent drug target for treating protistal diseases. Experimental screening to find drug candidates is an important step to discover new hit compounds. For that, a cheap, simple, and robust assay is needed. Here we report the application of the molybdenum blue reaction method for a medium throughput microplate activity assay of the hyperthermophilic bacterium Thermotoga maritima mPPase and the possible application of the assay to screen inhibitors of membrane-bound pyrophosphatases.
  • Hermans, Martijn; Lenstra, Wytze K.; van Helmond, Niels A. G. M.; Behrends, Thilo; Egger, Matthias; Seguret, Marie J. M.; Gustafsson, Erik; Gustafsson, Bo G.; Slomp, Caroline P. (2019)
    The Baltic Sea is characterized by the largest area of hypoxic (oxygen (O-2) <2 mg L-1) bottom waters in the world's ocean induced by human activities. Natural ventilation of these O-2-depleted waters largely depends on episodic Major Baltic Inflows from the adjacent North Sea. In 2014 and 2015, two such inflows led to a strong rise in O-2 and decline in phosphate (HPO42-) in waters below 125 m depth in the Eastern Gotland Basin. This provided the opportunity to assess the impact of such re-oxygenation events on the cycles of manganese (Mn), iron (Fe) and phosphorus (P) in the sediment for the first time. We demonstrate that the re-oxygenation induced the activity of sulphur (S)-oxidising bacteria, known as Beggiatoaceae in the surface sediment where a thin oxic and suboxic layer developed. At the two deepest sites, strong enrichments of total Mn and to a lesser extent Fe oxides and P were observed in this surface layer. A combination of sequential sediment extractions and synchrotron-based X-ray spectroscopy revealed evidence for the abundant presence of P-bearing rhodochrosite and Mn(II) phosphates. In contrast to what is typically assumed, the formation of Fe oxides in the surface sediment was limited. We attribute this lack of Fe oxide formation to the high flux of reductants, such as sulphide, from deeper sediments which allows Fe (II) in the form of FeS to be preserved and restricts the penetration of O-2 into the sediment. We estimate that enhanced P sequestration in surface sediments accounts for only similar to 5% of water column HPO42- removal in the Eastern Gotland Basin linked to the recent inflows. The remaining HPO42- was transported to adjacent areas in the Baltic Sea. Our results highlight that the benthic O-2 demand arising from the accumulation of organic-rich sediments over several decades, the legacy of hypoxia, has major implications for the biogeochemical response of euxinic basins to re-oxygenation. In particular, P sequestration in the sediment in association with Fe oxides is limited. This implies that artificial ventilation projects that aim at removing water column HPO42- and thereby improving water quality in the Baltic Sea will likely not have the desired effect. (C) 2018 Elsevier Ltd. All rights reserved.