Stereochemical probes of the argininosuccinate synthetase reaction.
The stereochemical course of the argininosuccinate synthetase reaction has been determined. The SP isomer of [alpha-17O,alpha-18O,alpha beta-18O]ATP is cleaved to (SP)-[16O,17O,18O]AMP by the action of argininosuccinate synthetase in the presence of citrulline and aspartate. The overall stereochemical transformation is therefore net inversion, and thus the enzyme does not catalyze the formation of an adenylylated enzyme intermediate prior to the synthesis of citrulline adenylate. The RP isomer of adenosine 5'-O-(2-thiotriphosphate) (ATP beta S) is a substrate in the presence of Mg2+, but the SP isomer is a substrate when Cd2+ is used as the activating divalent cation. Therefore, the lambda screw sense configuration of the beta,gamma-bidentate metal--ATP complex is preferred by the enzyme as the actual substrate. No significant discrimination could be detected between the RP and SP isomers of adenosine 5'-O-(1-thiotriphosphate) (ATP alpha S) when Mg2+ or Mn2+ are used as the divalent cation. Argininosuccinate synthetase has been shown to require a free divalent cation for full activity in addition to the metal ion needed to complex the ATP used in the reaction.
D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001124:Argininosuccinate Synthase / Q000378:metabolism*; D002417:Cattle; D007553:Isotope Labeling / Q000379:methods; D007700:Kinetics; D008025:Ligases / Q000378:metabolism*; D008099:Liver / Q000201:enzymology; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008968:Molecular Conformation; D010103:Oxygen Isotopes
D010103:Oxygen Isotopes; D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate; D008025:Ligases; D001124:Argininosuccinate Synthase