Adenine phosphoribosyl transferase: assay using high performance liquid chromatography

Abstract

Adenine phosphoribosyl transferase (APRT) (EC 2.4.2.7) is the enzyme which catalyses the reaction of the purine base adenine with S-phosphoribosyl-l-pyrophosphate (PP-ribose-P) to form the mononu~Ieotide adenosine monophosphate. Human APRT is a cytoplasmic enzyme present in all tissues with the highest specific activities being found in nucleated red cells. Severe homozygous deficiency of APRT is associated with nephrolithiasis and passage of urinary calculi composed of 2,8-dihydroxyadenine [1,2], suggesting that the enzyme has an important salvage function in man. Partial APRT deficiency [3,4] with 25-40% residual enzyme activity in red cell lysates is a relatively common heterozygous trait occurring in about 1% of the population [5] and is not associated with disease. Elevated levels of red cell APRT activity are found in the neonatal period [6] and in some patients with megalobtastic anaemia [7], as a direct consequence of reticulocytosis. They are also a characteristic finding in patients with Lesch-Nyhan syndrome [8] and the X-linked form of severe gout and hyperuricaemia associated with hypoxanthine-guanine phosphoribosyl transferase deficiency 191, where substrate stabilisation of PP-ribose-P may be important. Elevated erythrocyte APRT levels have also recently been reported in patients with hereditary erotic aciduria [lo], an inborn error of pyrimidine metabolism. Current methods for the quantitative assay of the enzyme are mainly radiochemical [ll-131. In this paper we describe an assay for red cell APRT activity utilising high performance liquid chromatography which is suitable for use in clinical laboratories,