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Metal drives chemistry: dual‐function of acireductone dioxygenase enzymes
Author(s) -
Deshpande Aditi Rajiva,
Wagenpfeil Karina,
Pochapsky Thomas Charles,
Petsko Gregory,
Ringe Dagmar
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.606.2
Subject(s) - chemistry , enzyme , formate , carbon monoxide , metal , catalysis , transition metal , stereochemistry , biochemistry , organic chemistry
Acireductone dioxygenase (ARD) from the methionine salvage pathway of Klebsiella oxytoca is the only known naturally occurring metalloenzyme which exhibits dual‐chemistry in vivo determined solely by the identity of the divalent transition metal ion (Fe 2+ or Ni 2+ ) in the active site. The Fe 2+ containing isozyme catalyzes the on‐pathway reaction using substrates acireductone and dioxygen to generate formate and the ketoacid precursor of methionine, whereas the Ni 2+ containing isozyme uses the same substrates to catalyze an off‐pathway shunt to methylthiopropionate, carbon monoxide and formate. We recently have shown that mammalian homologs of ARD (mouse and human) are also capable of this metal‐dependent dual‐chemistry in vitro . The possibility of this dual chemistry in mammals is interesting since carbon monoxide, one of the products of off‐pathway reaction, is an anti‐apoptotic molecule. In addition, several biochemical and genetic studies have indicated an inhibitory role of human ARD in cancer. Recombinant mouse and human ARD isozymes were expressed and purified to obtain a homogeneous single transition metal (Mn, Fe, Co, Ni) bound form of the enzymes. The Fe 2+ ‐bound ARD showed highest activity compared to the other metal‐bound forms of ARD. Similar to ARD from Klebsiella , both human and mouse ARD bound to Ni 2+ , Co 2+ or Mn 2+ catalyzed the off‐pathway reaction whereas Fe 2+ ‐bound ARD catalyzed the on‐pathway reaction of the methionine salvage pathway. Thermal stability indicated a large difference in melting temperatures of the proteins; with Ni 2+ ‐bound ARD being the most stable followed by Co 2+ and Fe 2+ and Mn 2+ being the least stable. Crystal structures of Ni 2+ ‐bound mouse ARD co‐crystallized with product and substrate analogs provided insight into metal coordination, substrate and product binding coordination and catalytic mechanism. Solution NMR data of Fe 2+ , Ni 2+ and Co 2+ ‐bound human ARD isozymes indicate that the metal center imparts significant structural differences to the overall protein structure. Support or Funding Information This work was supported in part by a grant from the National Institutes of Health (GM26788 to G.A.P. and D.R.)