Metabolism of threo-beta-fluoroaspartate by H4 cells. Inhibition of adenylosuccinate lyase by fluoro analogs of its substrates.
DL-threo-beta-Fluoroaspartate is a substrate for the two enzymes in de novo purine biosynthesis that use aspartate, namely 4-(N-succino)-5-aminoimidazole-4-carboxamide ribonucleotide (SAICAR) synthetase and adenylosuccinate synthetase. With both enzymes, Vmax with threo-beta-fluoroaspartate is about 50% of that observed with aspartate. The products of the two enzyme reactions, threo-beta-fluoro-SAICAR and threo-beta-fluoroadenylosuccinate, are inhibitors of adenylosuccinate lyase purified from rat skeletal muscle. In 20 mM phosphate buffer, pH 7.4, the KI values for threo-beta-fluoro-SAICAR are 5 and 3 microM and for threo-beta-fluoroadenylosuccinate are 3 and 1 microM, in the SAICAR and adenylosuccinate cleavage reactions, respectively. In 20 mM 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid buffer, pH 7.4, the KI values for threo-beta-fluoro-SAICAR are approximately 0.14 and 0.03 microM and for threo-beta-fluoroadenylosuccinate are approximately 0.05 and 0.015 microM, in the same two reactions, respectively. These KI values are one-half to one-hundredth of the Km values for SAICAR and adenylosuccinate, the two substrates of adenylosuccinate lyase. After an 8-h incubation with 45 microM threo-beta-fluoroaspartate, H4 cells contain 200-300 microM threo-beta-fluoro-SAICAR and 60-90 microM threo-beta-fluoroadenylosuccinate. These concentrations of fluoro analogs are sufficient to substantially inhibit adenylosuccinate lyase and hence the de novo synthesis of purines in H4 cells.
D000264:Adenylosuccinate Lyase / Q000037:antagonists & inhibitors*; D000265:Adenylosuccinate Synthase / Q000378:metabolism*; D000818:Animals; D001224:Aspartic Acid / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D002460:Cell Line; D007700:Kinetics; D008025:Ligases / Q000378:metabolism*; D008114:Liver Neoplasms, Experimental; D008190:Lyases / Q000037:antagonists & inhibitors*; D009132:Muscles / Q000201:enzymology; D010453:Peptide Synthases / Q000378:metabolism; D011687:Purines / Q000096:biosynthesis; D051381:Rats; D013329:Structure-Activity Relationship
D011687:Purines; D001224:Aspartic Acid; C023463:beta-fluoroaspartic acid; D008190:Lyases; D000264:Adenylosuccinate Lyase; D008025:Ligases; D010453:Peptide Synthases; C022379:phosphoribosylaminoimidazole-succinocarboxamide synthetase; D000265:Adenylosuccinate Synthase