Open AccessRadioimmune determination of hypoxanthine phosphoribosyltransferase crossreacting material in erythrocytes of Lesch-Nyhan patients.
Author(s) -
Gurdev S. Ghangas,
Gregory Milman
Publication year - 1975
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.72.10.4147
Subject(s) - hypoxanthine guanine phosphoribosyltransferase , hypoxanthine phosphoribosyltransferase , phosphoribosyltransferase , lesch–nyhan syndrome , hypoxanthine , biochemistry , radioimmunoassay , chemistry , enzyme , adenine phosphoribosyltransferase , microbiology and biotechnology , biology , mutant , purine , gene
We have developed a sensitive radioimmunoassay capable of detecting and quantitating 20 ng of hypoxanthine phosphoribosyltransferase (EC 2.4.2.8; IMP:pyrophosphate phosphoribosyltransferase) protein. For this assay, hypoxanthine phosphoribosyltransferase from human erythrocytes was iodinated with 125I under mild conditions using hydrogen peroxide and lactoperoxidase attached to Sepharose-4B. Antisera prepared against homogeneous human hypoxanthine phosphoribosyltransferase precipitates the iodinated enzyme as effectively as the unlabeled enzyme. The radioimmunoassay has been used to look for hypoxanthine phosphoribosyltransferase crossreacting material in hemolysates from sixteen different patients with a marked genetic deficiency of this enzyme characteristic of the Lesch-Nyhan syndrome. Fifteen hemolysates contained no detectable (less than 1% of normal) crossreacting material. One hemolysate contained a normal amount of crossreacting material. Hypoxanthine phosphoribosyltransferase from this patient (E.S.) has been shown to be a Km mutant enzyme.