Dual divalent cation requirement of the MutT dGTPase. Kinetic and magnetic resonance studies of the metal and substrate complexes.

MutT from the fish pathogenAliivibrio salmonicida is a cold‐active nucleotide‐pool sanitization enzyme with unexpectedly high thermostability

“…Although VcMutT was purified with 10 mM MgCl 2 , no electron density could be seen in the VcMutT structure corresponding to Mg 2+ . This is not a surprise knowing that the affinity of MutT to Mg 2+ is very low (1/50 compared to Mn 2+ ) [12]. However, when we model Mn 2+ in VcMutT-closed and AsMutT-closed assuming it is bound in the same sites as Mn in EcMutT, we observe that residues Glu54, Glu58, and the main-chain oxygen of Gly38 are potential metal binding residues compared to EcMutT (PDB: 3A6U) [10,35] thus we believe Mg 2+ can bind to our enzymes and thus contribute to the catalytic process (Fig.…”

“…MutT belongs to a superfamily of enzymes requiring two divalent cations for activity [12,13], and it has been shown that MutT possesses higher activity when the magnesium concentration is increased up to 10 mM [34]. Although VcMutT was purified with 10 mM MgCl 2 , no electron density could be seen in the VcMutT structure corresponding to Mg 2+ .…”

“…The MutT enzyme is widespread in nature and representatives are found in eukaryotes, prokaryotes, archaea and viruses [11]. It belongs to a superfamily of enzymes shown to require two divalent cations for activity [12,13]. Members of this family are recognised by a highly conserved 23-residue motif, or Nudix box, GX 5 EX 7 -REUXEEXGU, where U is a bulky hydrophobic or aliphatic residue and X is any residue [14,15].…”

Crystal Structure and Substrate Specificity of the 8-oxo-dGTP Hydrolase NUDT1 from Arabidopsis thaliana

“…This lack of activity can be explained by our 8-oxo-dGTP-bound structure, which shows that Glu56 contributes to the hydrogen bond network involved in coordinating the magnesium ions critical for the correct positioning and activation of the phosphate group. Extensive mechanistic studies of E. coli MutT have shown the enzyme requires two magnesium ions for activity. , These divalent cations exhibit octahedral coordination involving conserved glutamate residues and a glycine from the conserved Nudix motif, the phosphate group of the substrate, and additional water molecules . Catalysis occurs by nucleophilic substitution by water at the internal β-phosphate atom, generating the mononucleotide form of the substrate and pyrophosphate .…”

“…Extensive mechanistic studies of E. coli MutT have shown the enzyme requires two magnesium ions for activity. 60,61 These divalent cations exhibit octahedral coordination involving conserved glutamate residues and a glycine from the conserved Nudix motif, the phosphate group of the substrate, and additional water molecules. 62 Catalysis occurs by nucleophilic substitution by water at the internal β-phosphate atom, generating the mononucleotide form of the substrate and pyrophosphate.…”