title: Identification and purification of hydroxyisourate hydrolase, a novel ureide-metabolizing enzyme.
abstract: We report the identification and purification of a novel enzyme from soybean root nodules that catalyzes the hydrolysis of 5-hydroxyisourate, which is the true product of the urate oxidase reaction. The product of this reaction is 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline, and the new enzyme is designated 5-hydroxyisourate hydrolase. The enzyme was purified from crude extracts of soybean root nodules approximately 100-fold to apparent homogeneity with a final specific activity of 10 micromol/min/mg. The enzyme exhibited a native molecular mass of approximately 68 kDa by gel filtration chromatography and migrated as a single band on SDS-polyacrylamide gel electrophoresis with a subunit molecular mass of 68 +/- 2 kDa. The purified enzyme obeyed normal Michaelis-Menten kinetics, and the K(m) for 5-hydroxyisourate was determined to be 15 microM. The amino-terminal end of the purified protein was sequenced, and the resulting sequence was not found in any available data bases, confirming the novelty of the protein. These data suggest the existence of a hitherto unrecognized enzymatic pathway for the formation of allantoin.
authors.affiliation: Department of Biochemistry, University of Missouri, Columbia, Missouri 65211, USA.
authors.forename: A D
authors.initials: AD
authors.lastname: Sarma
authors.forename: P
authors.initials: P
authors.lastname: Serfozo
authors.forename: K
authors.initials: K
authors.lastname: Kahn
authors.forename: P A
authors.initials: PA
authors.lastname: Tipton
mesh_terms: D000595:Amino Acid Sequence; D006867:Hydrolases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D018517:Plant Roots / Q000201:enzymology; D017421:Sequence Analysis; D013025:Glycine max / Q000201:enzymology; D014508:Urea / Q000031:analogs & derivatives / Q000378:metabolism*
chemical_list: D014508:Urea; D006867:Hydrolases