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Metabolism and Enzymology

Ureide Catabolism in Soybeans ¹

III. Ureidoglycolate Amidohydrolase and Allantoate Amidohydrolase Are Activities of an Allantoate Degrading Enzyme Complex

Interdisciplinary Plant Biochemistry and Physiology Group, Curtis Hall, University of Missouri, Columbia, Missouri 65211

We demonstrate that allantoate is catabolized in soybean seedcoat extracts by an enzyme complex that has allantoate amidohydrolase and ureidoglycolate amidohydrolase activities. Soybean seedcoat extracts released ¹⁴CO₂ from [ureido-¹⁴C]ureidoglycolate under conditions in which urease is not detectable. CO₂ and glyoxylate are enzymically released in a one to one ratio indicating that ureidoglycolate amidohydrolase is the responsible activity. Ureidoglycolate amidohydrolase has a K_m of 85 micromolar for ureidoglycolate. Glyoxylate and CO₂ are enzymically released from allantoate at linear rates in a one to 2.3 ratio from 5 to 30 min. This ratio is consistent with the degradation of allantoate to two CO₂ and one glyoxylate with approximately 23% of the allantoate degraded reacting with 2-mercaptoethanol to yield 2-hydroxyethylthio, 2'-ureido, acetate (RG Winkler, JC Polacco, DG Blevins, DD Randall 1985 Plant Physiol 79: 787-793). That [¹⁴C]urea production from [2,7-¹⁴C]allantoate is not detectable indicates that allantoate-dependent glyoxylate production is enzymic and not a result of nonenzymic hydrolysis of a ureido intermediate (nonenzymic hydrolysis releases urea). These results and those from intact tissue studies (RG Winkler DG Blevins, JC Polacco, DD Randall 1987 Plant Physiol 83: 585-591) suggest that soybeans have a second amidohydrolase reaction (ureidoglycolate amidohydrolase) that follows allantoate amidohydrolase in allantoate catabolism. The rate of ¹⁴CO₂ release from [2,7-¹⁴C]allantoate is not reduced when the volume of the reaction mixture is increased, suggesting that the release of ¹⁴CO₂ is not dependent on the accumulation of free intermediates. That [2,7-¹⁴C]allantoate dependent ¹⁴CO₂ release is not proportionally diluted by unlabeled ureidoglycolate indicates that the reaction is carried out by an enzyme complex. This is the first report of ureidoglycolate amidohydrolase activity in any organism and the first in vitro demonstration in plants that the ureido-carbons of allantoate can be completely degraded to CO₂ without a urea intermediate.

¹ Supported by Missouri Agricultural Experiment Station and a grant from the United States Department of Agriculture, Science and Education Administration, Competitive Grants Office, Grant 85-CRCR-1-1638. This research is a contribution of the Missouri Agricultural Experiment Station Journal Series NO. 10357.

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