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A Purification and In Vitro Activity Assay for a (p)ppGpp Synthetase from Clostridium difficile
Published on: November 3, 2018
Structural characterization of the stringent response related exopolyphosphatase/guanosine pentaphosphate
Ole Kristensen1, Martin Laurberg, Anders Liljas
1Biostructural Research, Department of Medicinal Chemistry, The Danish University of Pharmaceutical Sciences, Universitetsparken 2, DK-2100 Copenhagen, Denmark. ok@dfuni.dk
Abstract:
Exopolyphosphatase/guanosine pentaphosphate phosphohydrolase (PPX/GPPA) enzymes play central roles in the bacterial stringent response induced by starvation. The high-resolution crystal structure of the putative Aquifex aeolicus PPX/GPPA phosphatase from the actin-like ATPase domain superfamily has been determined, providing the first insights to features of the common catalytic core of the PPX/GPPA family. The protein has a two-domain structure with an active site located in the interdomain cleft. Two crystal forms were investigated (type I and II) at resolutions of 1.53 and 2.15 A, respectively. This revealed a structural flexibility that has previously been described as a "butterfly-like" cleft opening around the active site in other actin-like superfamily proteins. A calcium ion is observed at the center of this region in type I crystals, substantiating that PPX/GPPA enzymes use metal ions for catalysis. Structural analysis suggests that nucleotides bind at a similar position to that seen in other members of the superfamily.
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