Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.4.2.7 (adenine phosphoribosyltransferase)
692 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Erythrocytes, obtained from a normal adult male and from a patient with Lesch-Nyhan syndrome, were incubated with [8-14C]adenine and [8-14C]hypoxanthine (Table 1). The labeled adenine was utilized to about the same extent for the synthesis of AMP by the normal subject's and the patient's erythrocytes. Deamination of AMP to IMP occurred to about the same extent in both samples. In contrast, hypoxanthine was utilized extensively for IMP synthesis in the normal erythrocyte only. The amount of total label in the IMP was about 100 times that of the Lesch-Nyhan erythrocyte, a consequence of the deficiency of hypoxanthine-guanine phosphoribosyltransferase (HGPRT) activity in the syndrome. No significant labeling of the AMP occurred. When aliquots of erythrocytes from both sources were incubated with 4-amino-5-imidazolecarboxamide (AICA) and sodium [14C]formate, extensive labeling of the IMP occurred in normal and in Lesch-Nyhan erythrocytes. The data suggest that AICA serves as a substrate for the adenine phosphoribosyltransferase (APRT) of the Lesch-Nyhan erythrocyte and that the ribotide of AICA, 5'-phosphoribosyl-5-aminoimidazole-4-carboxamide (AICAR), undergoes formylation by labeled N10-formyl tetrahydrofolic acid formed from the reaction of sodium [14C]formate with the tetrahydrofolic acid of the cell. The formyl-AICAR undergoes ring closure to IMP by a series of reactions comparable to those described for the normal erythrocyte. When 5-amino-1-ribosyl-4-imidazolecarboxamide (rAICA) and sodium [14C]formate were incubated with erythrocyte suspensions, extensive utilization for IMP synthesis was also observed in normal erythrocytes and in erythrocytes from Lesch-Nyhan patients (Table 2). The reaction sequence is somewhat different from that of AICA. AICA is not a substrate for the purine nucleoside phosphorylase of rabbit or human erythrocytes. The mechanism of rAICA utilization is visualized as a direct phosphorylation of the ribosyl compound, possibly by the adenosine kinase of the human cell. The ribotide, AICAR, formed by this mechanism, undergoes formylation and ring closure, yielding IMP. The glutamine antagonist, diazooxonorleucine (DON), was added to aliquots of patients' cells incubated with rAICA and sodium [14C]formate. DON is an effective inhibitor of the conversion of IMP to GMP and its presence in an incubation suspension resulted in a somewhat greater radioactivity of the total cellular IMP. The extension of the current studies to Lesch-Nyhan cells in culture may serve to assist in the direct evaluation of the regulatory role of IMP in the de novo pathway of purine nucleotide biosynthesis. Because of the substrate requirements of the reactions, the metabolism of AICA and rAICA may also serve to differentiate the roles of purine nucleotides and of phosphoribosylpyrophosphate (PRPP) in the pathway regulation. The findings presented also offer a possible therapeutic approach to the early treatment of the disease in the afflicted neonate...
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PMID:Lesch-Nyhan syndrome: the synthesis of inosine 5'-phosphate in the hypoxanthine-guanine phosphoribosyltransferase-deficient erythrocyte by alternate biochemical pathways. 87 Aug 76

Adenine and adenosine metabolism has been studied in intact human erythrocytes in vitro using high performance liquid chromatography, isotopic labeling and electrophoresis. Their metabolism to nucleotides was controlled by phosphoribose diphosphate synthesis which was phosphate dependent. Adenosine formed hypoxanthine or IMP depending upon Pi concentration, but adenosine kinase and deaminase activities were not affected by P levels. Free [14C]adenine and [14C]hypoxanthine were found in cellular extracts. Rapid interconversions occurred to give a distribution for ATP : ADP : AMP of 10 : 1 : 0.1. Marked decomposition of ATP to ADP and AMP occurred during incubations in plasma and Earle's media in air on nitrogen, but ATP levels remained stable in phosphate buffers and in the presence of oxygen. At physiological Pi (1 mM) adenosine kinase activity grossly exceeded adenine phosphoribosyltransferase activity. The latter was approximately 7 fold that of hypoxanthine phosphoribosyltransferase activity. These differences decreased with increasing Pi levels. No significant increase in corresponding nucleotides was obtained by incubation with high levels (0.5 mM) of adenine, guanine or guanosine at physiological Ii, ATP increased by 10% independently of the substrate employed and significant amounts of IMP and GTP were formed adenosine and guanosine, respectively. The existence of a bound intracellular pool of ATP is suggested.
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PMID:Studies on adenine and adenosine metabolism by intact human erythrocytes using high performance liquid chromatography. 94 98

8-Azainosine (8-aza-HR) is of interest because of its activity against experimental tumors. Metabolic studies in cell cultures were performed with 8-aza-HR and with the structurally related nucleoside, 8-azaadenosine (9-beta-D-ribofuranosyl-8-azaadenine) (8-aza-AR), which has a lower degree of antitumor activity than does 8-aza-HR. In H. Ep. 2 cells and in Ca755 cells, both 14C-labeled nucleosides were metabolized to nucleotides of 8-azaadenine (8-aza-A) and 8-azaguanine (8-aza-G) and incorporated into polynucleotides as 8-aza-A and 8-aza-G. 8-Aza HR was incorporated primarily as 8-aza-G, whereas 8-aza-AR was incorporated about equally as 8-aza-A and 8-aza-G. In H. Ep. 2 cells, the extent of incorporation of 8-aza-HR as 8-aza-G was about one-half that found when [14C]-8-aza-G was the precursor. In the H. Ep. 2/FA/FAR cell line, 8-aza-AR and 8-aza-HR were metabolized similarly, in that both were incorporated into polynucleotides principally as 8-aza-G; apparently, in this cell line which is deficient in adenosine kinase and adenine phosphoribosyltransferase, 8-aza-AR is metabolized by conversion to 8-aza-HR. A cell line (H. Ep 2/8-aza HR), which was resistant to 8-aza-HR but sensitive to 8-aza-AR and which retained hypoxanthine (guanine)-phosphoribosyltransferase activity, metabolized 8-aza-HR to only a small extent. However, in this cell-line, 8-aza-AR was more extensively metabolized and was incorporated primarily as 8-aza-A. The failure of these cells to convert 8-aza-AR or 8-aza-HR to 8-aza-G indicates that the basis for resistance may be a change in the substrate specificities of the enzymes of guanosine monophosphate synthesis such that these cells no longer effectively convert 8-azainosine monophosphate to 8-azaguanosine monophosphate. 8-Aza-AR was a potent inhibitor of purine synthesis de novo, but 8-aza-HR, at concentrations much higher than the inhibitory concentration of 8-aza-AR, did not inhibit this process. In H. Ep. 2 cells, 8-aza-HR blocked the conversion of orotic acid to uridine nucleotides and caused an accumulation of orotidine. This inhibition of pyrimidine biosynthesis apparently does not contribute significantly to the cytotoxicity of 8-aza-HR because uridine provided no degree of reversal of its inhibition of the growth of cell cultures.
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PMID:Metabolism and metabolic effects of 8-azainosine and 8-azaadenosine. 97 40

To evaluate the regulation of adenine nucleotide metabolism in relation to purine enzyme activities in rat liver, human erythrocytes and cultured human skin fibroblasts, rapid and sensitive assays for the purine enzymes, adenosine deaminase (EC 2.5.4.4), adenosine kinase (EC 2.7.1.20), hyposanthine phosphoribosyltransferase (EC 2.4.28), adenine phosphoribosyltransferase (EC 2.4.2.7) and 5'-nucleotidase (EC 3.1.3.5) were standardized for these tissues. Adenosine deaminase was assayed by measuring the formation of product, inosine (plus traces of hypoxanthine), isolated chromatographically with 95% recovery of inosine. The other enzymes were assayed by isolating the labelled product or substrate nucleotides as lanthanum salts. Fibroblast enzymes were assayed using thin-layer chromatographic procedures because the high levels of 5'-nucleotidase present in this tissue interferred with the formation of LaCl3 salts. The lanthanum and the thin-layer chromatographic methods agreed within 10%. Liver cell sap had the highest activities of all purine enzymes except for 5'-nucleotidase and adenosine deaminase which were highest in fibroblasts. Erythrocytes had lowest activities of all except for hypoxanthine phosphoribosyltransferase which was intermediate between the liver and fibroblasts. Erhthrocytes were devoid of 5'-nucleotidase activity. Hepatic adenosine kinase activity was thought to control the rate of loss of adenine nucleotides in the tissue. Erythrocytes had excellent purine salvage capacity, but due to the relatively low activity of adenosine deaminase, deamination might be rate limiting in the formation of guanine nucleotides. Fibroblasts, with high levels of 5'-nucleotidase, have the potential to catabolize adenine nucleotides beyond the control od adenosine kinase. The purine salvage capacity in the three tissues was erythrocyte greater than liver greater than fibroblasts. Based on purine enzyme activities, erythrocytes offer a unique system to study adenine salvage; fibroblasts to study adenine degradation; and liver to study both salvage and degradation.
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PMID:Adenine nucleotide metabolism in relation to purine enzymes in liver, erythrocytes and cultured fibroblasts. 118 98

Adenosine derivatives are frequently used in chemotherapy because of their potent antitumor, antiviral and antiparasitic activity. We investigated the metabolism of some adenosine analogues in adenosine deaminase inhibited normal and adenine phosphoribosyltransferase (APRT) deficient human erythrocytes. The ATP and GTP concentrations and the formation of unusual nucleotides were measured. Some of the analogues studied (tubercidin, 9 beta-D-arabinofuranosyladenine, 2'-deoxyadenosine, 2-chloroadenosine, neplanocin A) were phosphorylated to the corresponding nucleoside triphosphates and this process was abolished by iodotubercidin--an adenosine kinase inhibitor. With the exception of 2'-deoxyadenosine, nucleotide analogue formation was accompanied by ATP depletion. ATP decrease was not observed after adenosine kinase inhibition and ATP concentration even increased in the presence of 2'-deoxyadenosine, neplanocin A and 5'-iodo-5'-deoxyadenosine. However, the latter increment was not observed in APRT deficient erythrocytes. Bredinin, S-adenosylhomocysteine, deoxycoformycin and adenosine dialdehyde did not form nucleotide derivatives or exert any effects on ATP concentration. It is concluded that adenosine analogues can either enter the nucleotide pool via phosphorylation mechanisms, or may be converted to ATP by the pathways involving the intermediate formation of adenine.
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PMID:Effects of adenosine analogues on ATP concentrations in human erythrocytes. Further evidence for a route independent of adenosine kinase. 193 Mar 1

The aim of this study was to identify targets for rational chemotherapy of glioblastoma. In order to elucidate differences in the biochemistry of tumor and normal human brain, in vivo pool sizes of purine nucleotides, nucleosides, and nucleobases and of purine metabolizing enzymes in biopsy material from 14 grade IV astrocytomas and 4 normal temporal lobe samples were analyzed. Specimens were collected during surgery using the freeze-clamp sampling technique and analyzed by high pressure liquid chromatography. Total purine nucleotides, adenylates, and guanylates in the tumors were 2186, 1865, and 310 nmol/g (wet weight), respectively, which corresponds to 61, 60, and 71% of normal brain tissue concentrations. Relative to normal brain the tumors had significantly lower ATP and GTP levels, essentially normal pool sizes of purine nucleosides and bases, unchanged activities of the salvage enzymes hypoxanthine-guanine phosphoribosyltransferase, adenine phosphoribosyltransferase, and adenosine kinase (659, 456, and 98 nmol/h/mg protein, respectively) and 4-fold higher activities of IMP dehydrogenase (11.6 nmol/h/mg protein); the latter is the rate limiting enzyme for guanylate de novo synthesis. IMP pools in the tumors were 64% of values in normal brain. Modulation of the guanylate pathway in glioblastoma by inhibition of IMP dehydrogenase with tumor specific agents such as tiazofurin (2-beta-D-ribofuranosylthiazole-4-carboxamide) appears to be a rational therapeutic approach. Preliminary in vitro experiments with normal and malignant tissue specimens from 2 additional patients revealed that significant amounts of the active metabolite thiazole-4-carboxamide adenine dinucleotide are formed from tiazofurin. At a concentration of 200 microM this drug was able to deplete guanylate pools in the tumors to a median of 54% of phosphate buffered saline treated controls. Flux studies with [14C]formate showed that tiazofurin strongly inhibited de novo synthesis of guanylates in glioblastoma to an average of 10% of controls. This effect was more pronounced in the tumors as compared to normal brain. No inhibition of salvage of [14C]guanine by tiazofurin could be observed in normal and malignant tissues. Supportive measures have to be considered to inhibit the highly active salvage enzyme hypoxanthine-guanine phosphoribosyltransferase that can partly antagonize a tiazofurin induced decrease in guanine nucleotides.
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PMID:Purine metabolism of human glioblastoma in vivo. 215 28

The mechanism by which S-adenosylmethionine (SAM) and adenosine (Ado) increase ATP levels in intact human erythrocytes in vitro has been compared. The use of erythrocytes from healthy controls and from subjects totally deficient in adenine phosphoribosyltransferase (APRT), plus inhibitors of adenosine kinase (AK) and adenosine deaminase (ADA) separately and together, has enabled us to demonstrate that this increment in ATP levels occurred via totally different metabolic routes. The results show that: (i) whilst the Ado-induced increment in ATP was AK dependent, that produced by SAM was independent of AK: and (ii) the SAM-induced increment in ATP was totally dependent on APRT and that some of the increment produced by Ado might also be APRT dependent. The above data are consistent with the metabolism of SAM to ATP by a route recently identified by us whereby ATP is formed from deoxyadenosine: namely binding to the enzyme S-adenosylhomocysteine hydrolase with subsequent release of adenine and further conversion to ATP via APRT.
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PMID:S-adenosylmethionine increases erythrocyte ATP in vitro by a route independent of adenosine kinase. 226 Sep 86

The activities (Vmax) of several enzymes of purine nucleotide metabolism were assayed in premature and mature primary rat neuronal cultures and in whole rat brains. In the neuronal cultures, representing 90% pure neurons, maturation (up to 14 days in culture) resulted in an increase in the activities of guanine deaminase (guanase), purine-nucleoside phosphorylase (PNP), IMP 5'-nucleotidase, adenine phosphoribosyltransferase (APRT), and AMP deaminase, but in no change in the activities of hypoxanthine-guanine phosphoribosyltransferase (HGPRT), adenosine deaminase, adenosine kinase, and AMP 5'-nucleotidase. In whole brains in vivo, maturation (from 18 days of gestation to 14 days post partum) was associated with an increase in the activities of guanase, PNP, IMP 5'-nucleotidase, AMP deaminase, and HGPRT, a decrease in the activities of adenosine deaminase and IMP dehydrogenase, and no change in the activities of APRT, AMP 5'-nucleotidase, and adenosine kinase. The profound changes in purine metabolism, which occur with maturation of the neuronal cells in primary cultures in vitro and in whole brains in vivo, create an advantage for AMP degradation by deamination, rather than by dephosphorylation, and for guanine degradation to xanthine over its reutilization for synthesis of GMP. The physiological meaning of the maturational increase in these two ammonia-producing enzymes in the brain is not yet clear. The striking similarity in the alterations of enzyme activities in the two systems indicates that the primary culture system may serve as an appropriate model for the study of purine metabolism in brain.
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PMID:Developmental changes in the activity of enzymes of purine metabolism in rat neuronal cells in culture and in whole brain. 232 47

1. Both normal cells and cells deficient in hypoxanthine-guanine phosphoribosyltransferase (HPRT) are able to produce adenine and guanine nucleotides from aminoimidazole carboxamide (AICA) or its ribonucleoside (AICAR), but not from formaminoimidazole carboxamide ribonucleoside (FAICAR). 2. The level of purine nucleotide production from AICA in HPRT- cells is at least equal to the production of purine nucleotides from hypoxanthine in normal cells. 3. The concentration of AICA or AICAR at which nucleotide production was half-maximal was between 30 and 100 microM in various cell lines. 4. Adenosine kinase is required to convert AICAR to its nucleotide; adenine phosphoribosyltransferase is required to convert AICA to its nucleotide. Cells lacking either of these enzymes are unable to produce purine nucleotides from the respective precursor. 5. Purine production from AICAR in HPRT- cells is not greatly increased by the addition of formate, folate or leucovorin.
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PMID:Purine nucleotide production in normal and HPRT- cells. 261 26

Using radiochemical methods, we determined the activities of various enzymes of purine and pyrimidine metabolism in homogenates of human skeletal muscle and of cultured human muscle cells. Results show a large discrepancy between the enzyme activities in muscle and cultured cells. With regard to purine metabolism, adenylate (AMP) deaminase activity was only 1-3% in cultured cells compared to that in muscle, whereas the activity of adenosine deaminase, purine-nucleoside phosphorylase, adenosine kinase, adenine phosphoribosyltransferase and hypoxanthine phosphoribosyltransferase was 7-15-fold higher in the cultured cells. The enzymes of pyrimidine metabolism, orotate phosphoribosyltransferase, orotidine 5'-monophosphate decarboxylase and uridine kinase showed activity of 100-200-fold higher in cultured cells than in adult muscle. The differences in enzyme activity are probably related to the low differentiation stage and the absence of contractile activity in the cultured muscle cells. Care must be taken when using these cells as a model for studying purine and pyrimidine metabolism of adult myofibers.
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PMID:Purine and pyrimidine metabolism in human muscle and cultured muscle cells. 283 95


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