J. Frank Henderson

Hypoxanthine-Guanine Phosphoribosyltransferase Deficiency in Gout

Excerpt INTRODUCTION Sophisticated biochemical studies in recent years have revealed that the regulation of intracellular metabolism is a logical, orderly, and intricate process. Control of enzyme ...

Adenine phosphoribosyltransferase deficiency: a previously undescribed genetic defect in man

A deficiency of adenine phosphoribosyltransferase (A-PRTase) is described in four members in three generations of one family. A-PRTase is coded by an autosome and the mutants described in this report are heterozygotes for this enzyme defect. The level of enzyme activity in these heterozygotes was inappropriately low, ranging from 21 to 37% of normal rather than the expected 50% of normal. Examination of various physical and chemical properties of the A-PRTase obtained from the mutant heterozygotes failed to reveal differences from the normal enzyme. These patients have no discernable abnormality in uric acid production despite the finding that patients with a deficiency of a closely related enzyme, hypoxanthine-guanine phosphoribosyltransferase, invariably produce excessive quantities of uric acid. A relationship of the A-PRTase deficiency to the disturbance in lipoprotein metabolism observed in the propositus has not been firmly established. Possible manifestations of the homozygous form of this enzyme deficiency will require identification of such individuals in the future.

Variations in purine metabolism of cultured skin fibroblasts from patients with gout

Purine metabolism was studied in fibroblasts cultured from three patients with gout in an attempt to determine the biochemical bases of their disease. The rate of purine biosynthesis de novo was normal in one line of cells, but the rate of catabolism of adenine nucleotides to hypoxanthine and inosine was greatly increased. The rate of purine biosynthesis de novo was increased in two lines of cells, and this was associated with increased concentrations of 5-phosphoribosyl 1-pyrophosphate. Purine synthesis was also less sensitive than normal to feedback inhibition. The catabolism of inosinate synthesized de novo was increased.

Chapter 19 Isolation of Drug-Resistant Clones of Ehrlich Ascites Tumor Cells

Publisher Summary Drug resistance is a continuing obstacle to effective cancer chemotherapy, and several experimental models have been used in attempts to understand, and counteract or circumvent this hindrance. These model systems fall into two major classes: selection of resistant clones from cells growing in culture, and the gradual enrichment in resistant cells of an ascites tumor population by in vivo selection for many weeks. The rate of spontaneous mutation to drug resistance in animal cell systems is relatively high, and it is important to clone resistant cell lines to ensure that the population is isogeneic. Although this is easily accomplished in the case of resistant cell lines isolated in vitro, resistant ascitic tumors isolated in vivo rarely have been cloned, and this is difficult to do. Although selection pressures used to isolate resistant ascites tumors are limited by host toxicity, relatively high drug does can usually be given because drugs administered intraperitoneally are often less toxic when an ascites tumor is present in the peritoneal cavity, and because the tumor can be transplanted to new hosts at relatively frequent intervals. Experimental studies of drug resistance relevant to the clinical situation have suffered from a lack of suitable model systems. The procedure described in this chapter was developed for the isolation of drug resistant clones of tumor cells in vivo using low selection pressures.

Hypoxanthine-guanine phosphoribosyltransferase: further evidence for the identity of the binding sites for hypoxanthine and guanine.

Isotope exchange between hypoxanthine and both inosinate and guanylate, and between guanine and the same two ribonucleotides, support the view that hypoxanthine and guanine bind to the same site on hypoxanthine-guanine phosphoribosyltransferase.

Altered Purine Metabolism in Regulatory Mutants of Saccharomyces cerevisiae

Armitt & Woods (1970) and Lomax & Woods (1973) reported the isolation of a series of mutants of Saccharomyces cerevisiae that did not require purines for growth, but excreted substantial amounts of purines into the growth medium. Genetic studies have assigned these yeast strains to six unlinked genes (purl, pur2, pur3, pur4, pur.5, pzv6), of which one, pur6, had both recessive and dominant (PUR6) alleles. Purine excretion was prevented by another gene, su-pur, except in an allele of purl called purI(ps), in which only partial suppression took place. Mapping revealed that pur6 was allelic to ade4, which has been identified (Gross & Woods, 197 1) as the structural gene for amidophosphoribosyltransferase EC 2.4.2.14), the first enzyme of the pathway of purine biosynthesis de novo. Purl was allelic to adel2 (Lomax & Woods, 1971), the gene for adenylosuccinate synthetase (EC 6.3.4.4); this locus is in fact bifunctional, and in addition to the catalysis of the adenylosuccinate synthetase reaction it also regulates the rate of the de novo purine biosynthetic pathway (Dorfman, 1965, 1971; Lomax & Woods, 1971). There is also evidence that the pur5 locus is involved in the regulation of the synthesis or function of inosinate dehydrogenase (EC 1.2.1.14) (W. J. E. Gardner & R. W. Woods, unpublished observations). Because the pur mutants excrete purines into the medium at considerable rates but can depend entirely on purine biosynthesis de novo to supply purines for growth, it has been assumed that they have accelerated rates of purine biosynthesis de novo and hence that they are in some way altered in the regulation of this pathway. The excretion of purines, mostly hypoxanthine and inosine (Lomax & Woods, 1973), into the medium has therefore been thought to be a direct consequence of the accelerated purine biosynthesis de novo, and hence a way of disposing of unneeded inosinate. In order to test these hypotheses, and to elucidate other aspects of purine metabolism in these yeast strains, we have made quantitative studies of the metabolism of radioactive glycine, adenine, guanine and hypoxanthine.

CLASSIFICATION OF PURINE NUCLEOSIDE ANALOGS BASED ON MULTIPLE BIOLOGICAL AND BIOCHEMICAL PARAMETERS: 145

On the basis of a systematic study of the effects of growth inhibitory purine and purine nucleoside analogs on 29 biological and biochemical parameters, we have classified them into 12 major groups. Drugs whose toxicity is reversed by adenine and hypoxanthine are divided into 3 groups. In addition to reduction of formate incorporation, altered ribonucleotide pool sizes and inhibition of macromolecule synthesis, 1 group is metabolized to drug nucleotides whereas the 2nd group is not. The 3rd group has diverse effects. Drugs whose toxicity is not reversed by adenine and hypoxanthine and which demonstrate reduction of incorporation into nucleotides of both formate and hypoxanthine are sub-divided into 4 categories. The first 2 inhibit macro-molecule synthesis with differences in drug nucleotide formation and cell volume. The latter 2 categories form drug nucleotides and show diverse effects on ribonucleotide pools and DNA, RNA and protein synthesis. Five groups remain. One markedly inhibits DNA synthesis, a 2nd involves IMP dehydrogenase inhibition, and a 3rd shows high GTP levels and marked reduction of formate and hypoxanthine incorporation. The last 2 groups, involving potentiation by deoxycoformycin or guanine, overlap with the groups already discussed. This study directs attention to promising analogs which warrant further biochemical and pharmacological investigation and perhaps clinical trial.