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Purification and characterization of recombinant human liver glycolate oxidase.

Abstract
Glycolate oxidase, an FMN-dependent peroxisomal oxidase, plays an important role in plants, related to photorespiration, and in animals, where it can contribute to the production of oxalate with formation of kidney stones. The best studied plant glycolate oxidase is that of spinach; it has been expressed as a recombinant enzyme, and its crystal structure is known. With respect to animals, the enzyme purified from pig liver has been characterized in detail in terms of activity and inhibition, the enzyme from human liver in less detail. We describe here the purification and initial characterization of the recombinant human glycolate oxidase. Its substrate specificity and the inhibitory effects of a number of anions are in agreement with the properties expected from previous work on glycolate oxidases from diverse sources. The recombinant enzyme presents an inhibition by excess glycolate and by excess DCIP, which has not been documented before. These inhibitions suggest that glycolate binds to the active site of the reduced enzyme, and that DCIP also has affinity for the oxidized enzyme. Glycolate oxidase belongs to a family of l-2-hydroxy-acid-oxidizing flavoenzymes, with strongly conserved active-site residues. A comparison of some of the present results with studies dealing with other family members suggests that residues outside the active site influence the binding of a number of ligands, in particular sulfite.
AuthorsCaroline Vignaud, Nicolas Pietrancosta, Emma L Williams, Gill Rumsby, Florence Lederer
JournalArchives of biochemistry and biophysics (Arch Biochem Biophys) Vol. 465 Issue 2 Pg. 410-6 (Sep 15 2007) ISSN: 0003-9861 [Print] United States
PMID17669354 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
Chemical References
  • Recombinant Proteins
  • Alcohol Oxidoreductases
  • glycollate oxidase
Topics
  • Alcohol Oxidoreductases (chemistry, genetics, isolation & purification, metabolism)
  • Enzyme Activation
  • Enzyme Stability
  • Humans
  • Liver (enzymology)
  • Recombinant Proteins (chemistry, isolation & purification, metabolism)
  • Substrate Specificity

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