Evidence for the Direct Transfer of the Carboxylate of N super(5)-Carboxyaminoimidazole Ribonucleotide (N super(5)-CAIR) To Generate 4-Carboxy-5-aminoimidazole Ribonucleotide Catalyzed by Escherichia coli PurE, an N super(5)-CAIR Mutase

• Published in: Biochemistry (Easton) 1999-03, Vol.38 (10), p.3012-3018
• Main Authors: Meyer, E, Kappock, T J, Osuji, C, Stubbe, J
• Format: Article
• Language: English
• Subjects: 4-carboxy-5-aminoimidazole ribonucleotide; 5-aminoimidazole ribonucleotide; carboxylic acids; Escherichia coli; PurE protein; PurK protein; ribonucleotide synthase
• ISSN: 0006-2960
• DOI: 10.1021/bi9827159

Formation of 4-carboxy-5-aminoimidazole ribonucleotide (CAIR) in the purine pathway in most prokaryotes requires ATP, HCO sub(3) super(-), aminoimidazole ribonucleotide (AIR), and the gene products PurK and PurE. PurK catalyzes the conversion of AIR to N super(5)-carboxyaminoimidazole ribonucleotide (N super(5)-CAIR) in a reaction that requires both ATP and HCO sub(3) super(-). PurE catalyzes the unusual rearrangement of N super(5)-CAIR to CAIR. To investigate the mechanism of this rearrangement, [4,7- super(13)C]-N super(5)-CAIR and [7- super(14)C]-N super(5)-CAIR were synthesized and separately incubated with PurE in the presence of ATP, aspartate, and 4-(N-succinocarboxamide)-5-aminoimidazole ribonucleotide (SAICAR) synthetase (PurC). The SAICAR produced was isolated and analyzed by NMR spectroscopy or scintillation counting, respectively. The PurC trapping of CAIR as SAICAR was required because of the reversibility of the PurE reaction. Results from both experiments reveal that the carboxylate group of the carbamate of N super(5)-CAIR is transferred directly to generate CAIR without equilibration with CO sub(2)/HCO sub(3) super(-) in solution. The mechanistic implications of these results relative to the PurE-only (CO sub(2)- and AIR-requiring) AIR carboxylases are discussed.