Pentatrichomonas hominis: purine salvage pathway

1. Pentatrichomonas hominis was found incapable of de novo synthesis of purines. 2. Pentatrichomonas hominis can salvage adenine, guanine, hypoxanthine, adenosine, guanosine and inosine, but not xanthine for the synthesis of nucleotides. 3. HPLC tracing of radiolabelled purines or purine nucleosides revealed that adenine, adenosine and hypoxanthine are incorporated into adenine nucleotides and IMP through a similar channel while guanine and guanosine are salvaged into guanine nucleotides via another route. There appears to be no direct interconversion between adenine and guanine nucleotides. Interconversion between AMP and IMP was observed. 4. Assays of purine salvage enzymes revealed that P. hominis possess adenosine kinase; adenosine, guanosine and inosine phosphotransferases; adenosine, guanosine and inosine phosphorylases and AMP deaminase.     

Profiles of purine biosynthesis, salvage and degradation in disks of potato (Solanum tuberosum L.) tubers

To find general metabolic profiles of purine ribo- and deoxyribonucleotides in potato (Solanum tuberosum L.) plants, we looked at the in situ metabolic fate of various ¹⁴C-labelled precursors in disks from growing potato tubers. The activities of key enzymes in potato tuber extracts were also studied. Of the precursors for the intermediates in de novo purine biosynthesis, [¹⁴C]formate, [2-¹⁴C]glycine and [2-¹⁴C]5-aminoimidazole-4-carboxyamide ribonucleoside were metabolised to purine nucleotides and were incorporated into nucleic acids. The rates of uptake of purine ribo- and deoxyribonucleosides by the disks were in the following order: deoxyadenosine > adenosine > adenine > guanine > guanosine > deoxyguanosine > inosine > hypoxanthine > xanthine > xanthosine. The purine ribonucleosides, adenosine and guanosine, were salvaged exclusively to nucleotides, by adenosine kinase (EC 2.7.1.20) and inosine/guanosine kinase (EC 2.7.1.73) and non-specific nucleoside phosphotransferase (EC 2.7.1.77). Inosine was also salvaged by inosine/guanosine kinase, but to a lesser extent. In contrast, no xanthosine was salvaged. Deoxyadenosine and deoxyguanosine, was efficiently salvaged by deoxyadenosine kinase (EC 2.7.1.76) and deoxyguanosine kinase (EC 2.7.1.113) and/or non-specific nucleoside phosphotransferase (EC 2.7.1.77). Of the purine bases, adenine, guanine and hypoxanthine but not xanthine were salvaged for nucleotide synthesis. Since purine nucleoside phosphorylase (EC 2.4.2.1) activity was not detected, adenine phosphoribosyltransferase (EC 2.4.2.7) and hypoxanthine/guanine phosphoribosyltransferase (EC 2.4.2.8) seem to play the major role in salvage of adenine, guanine and hypoxanthine. Xanthine was catabolised by the oxidative purine degradation pathway via allantoin. Activity of the purine-metabolising enzymes observed in other organisms, such as purine nucleoside phosphorylase (EC 2.4.2.1), xanthine phosphoribosyltransferase (EC 2.4.2.22), adenine deaminase (EC 3.5.4.2), adenosine deaminase (EC 3.5.4.4) and guanine deaminase (EC 3.5.4.3), were not detected in potato tuber extracts. These results suggest that the major catabolic pathways of adenine and guanine nucleotides are AMP → IMP → inosine → hypoxanthine → xanthine and GMP → guanosine → xanthosine → xanthine pathways, respectively. Catabolites before xanthosine and xanthine can be utilised in salvage pathways for nucleotide biosynthesis.     

Profiles of purine metabolism in leaves and roots of Camellia sinensis seedlings

To determine the metabolic profiles of purine nucleotides and related compounds in leaves and roots of tea (Camellia sinensis), we studied the in situ metabolic fate of 10 different (14)C-labeled precursors in segments from tea seedlings. The activities of key enzymes in tea leaf extracts were also investigated. The rates of uptake of purine precursors were greater in leaf segments than in root segments. Adenine and adenosine were taken up more rapidly than other purine bases and nucleosides. Xanthosine was slowest. Some adenosine, guanosine and inosine was converted to nucleotides by adenosine kinase and inosine/guanosine kinase, but these compounds were easily hydrolyzed, and adenine, guanine and hypoxanthine were generated. These purine bases were salvaged by adenine phosphoribosyltransferase and hypoxanthine/guanine phosphoribosyltransferase. Salvage activity of adenine and adenosine was high, and they were converted exclusively to nucleotides. Inosine and hypoxanthine were salvaged to a lesser extent. In situ (14)C-tracer experiments revealed that xanthosine and xanthine were not salvaged, although xanthine phosphoribosyltransferase activity was found in tea extracts. Only some deoxyadenosine and deoxyguanosine was salvaged and utilized for DNA synthesis. However, most of these deoxynucleosides were hydrolyzed to adenine and guanine and then utilized for RNA synthesis. Purine alkaloid biosynthesis in leaves is much greater than in roots. In situ experiments indicate that adenosine, adenine, guanosine, guanine and inosine are better precursors than xanthosine, which is a direct precursor of a major pathway of caffeine biosynthesis. Based on these results, possible routes of purine metabolism are discussed.     

Genetic and physiological characterization of the purine salvage pathway in the archaebacterium Methanobacterium thermoautotrophicum Marburg.

The enzymes involved in the purine interconversion pathway of wild-type and purine analog-resistant strains of Methanobacterium thermoautotrophicum Marburg were assayed by radiometric and spectrophotometric methods. Wild-type cells incorporated labeled adenine, guanine, and hypoxanthine, whereas mutant strains varied in their ability to incorporate these bases. Adenine, guanine, hypoxanthine, and xanthine were activated by phosphoribosyltransferase activities present in wild-type cell extracts. Some mutant strains simultaneously lost the ability to convert both guanine and hypoxanthine to the respective nucleotide, suggesting that the same enzyme activates both bases. Adenosine, guanosine, and inosine phosphorylase activities were detected for the conversion of base to nucleoside. Adenine deaminase activity was detected at low levels. Guanine deaminase activity was not detected. Nucleoside kinase activities for the conversion of adenosine, guanosine, and inosine to the respective nucleotides were detected by a new assay. The nucleotide-interconverting enzymes AMP deaminase, succinyl-AMP synthetase, succinyl-AMP lyase, IMP dehydrogenase, and GMP synthetase were present in extracts; GMP reductase was not detected. The results indicate that this autotrophic methanogen has a complex system for the utilization of exogenous purines.     

Development of a FIA system with immobilized enzymes for specific post-column detection of purine bases and their nucleosides separated by HPLC column.

A sensitive and highly selective method for the simultaneous determination of purine bases and their nucleosides is proposed. An amperometric flow-injection system with the two immobilized enzyme reactors (guanase immobilized reactor and purine nucleoside phosphorylase/xanthine oxidase co-immobilized reactor) is used as the specific post-column detection system of HPLC, to convert compounds separated by a reversed-phase. HPLC column to electroactive species (hydrogen peroxide and uric acid) which can be detected at a flow-through platinum electrode. The proposed detection system is specific for a group of purine bases and purine nucleosides and does not respond for purine nucleotides and pyrimidine bases. The linear determination ranges are from 10 pmol to 5 nmol for four purine bases (hypoxanthine, xanthine, guanine, and adenine) and four purine nucleosides (inosine, xanthosine, guanosine, and adenosine). The detection limits are 1.2-5.5 pmol.     

Purine metabolism in Acholeplasma laidlawii B: novel PPi-dependent nucleoside kinase activity

Acholeplasma laidlawii B-PG9 was examined for 16 cytoplasmic enzymes with activity for purine salvage and interconversion. Phosphoribosyltransferase activities for adenine, guanine, xanthine, and hypoxanthine were shown. Adenine, guanine, xanthine, and hypoxanthine were ribosylated to their nucleoside. Adenosine, inosine, xanthosine, and guanosine were converted to their base. No ATP-dependent phosphorylation of nucleosides to mononucleotides was found. However, PPi-dependent phosphorylation of adenosine, inosine, and guanosine to AMP, inosine monophosphate, and GMP, respectively, was detected. Nucleotidase activity for AMP, inosine monophosphate, xanthosine monophosphate, and GMP was also found. Interconversion of GMP to AMP was detected. Enzyme activities for the interconversion of AMP to GMP were not detected. Therefore, A. laidlawii B-PG9 cannot synthesize guanylates from adenylates or inosinates. De novo synthesis of purines was not detected. This study demonstrates that A. laidlawii B-PG9 has the enzyme activities for the salvage and limited interconversion of purines and, except for purine nucleoside kinase activity, is similar to Mycoplasma mycoides subsp. mycoides. This is the first report of a PPi-dependent nucleoside kinase activity in any organism.     

Purine salvage in Entamoeba histolytica

Pulse-labeling of the nucleotide pool in Entamoeba histolytica with radioactive precursors, and subsequent high performance liquid chromatographic (HPLC) analysis of the radiolabeled nucleotides, indicate that E. histolytica is incapable of de novo synthesis of purine nucleotides. Hypoxanthine, inosine and xanthine could not be converted to nucleotides in E. histolytica, which suggests the absence of interconversion between adenine nucleotides and guanine nucleotides through formation of IMP. Adenosine was actively incorporated into nucleotides at an initial rate of 130 pmoles per minute per 10(6) trophozoites. Adenine, guanosine and guanine were also incorporated at much lower rates. The rate of adenine incorporation was enhanced by the presence of guanosine; the rate of guanine incorporation was significantly increased by adenosine. These stimulatory effects suggest that the ribose moiety of adenosine or guanosine can be transferred to another purine base to form a new nucleoside, and that the purine nucleosides are the immediate precursors of E. histolytica nucleotides. HPLC results showed that the radiolabel in adenine was exclusively incorporated into adenine nucleotides and that guanine was found only among guanine nucleotides, whereas the radioactivity associated with the ribose moiety of adenosine or guanosine was distributed among both adenine and guanine nucleotides.     

A comparison of purine metabolism and nucleotide pools in normal and hypoxanthine-guanine phosphoribosyltransferase-deficient neuroblastoma cells

Purine nucleotide synthesis and interconversion were examined over a range of purine base and nucleoside concentrations in intact N4 and N4TG (hypoxanthine-guanine phosphoribosyltransferase (HGPRT) deficient) neuroblastoma cells. Adenosine was a better nucleotide precursor than adenine, hypoxanthine or guanine at concentrations greater than 100 μM. With hypoxanthine or guanine, N4TG cells had less than 2% the rate of nucleotide synthesis of N4 cells. At substrate concentrations greater than 100 μM the rates for deamination of adenosine and phosphorolysis of guanosine exceeded those for any reaction of nucleotide synthesis. Labelled inosine and guanosine accumulated from hypoxanthine and guanine, respectively, in HGPRT-deficient cells and the nucleosides accumulated to a greater extent in N4 cells indicating dephosphorylation of newly synthesized IMP and GMP to be quantitatively significant. A deficiency of xanthine oxidase, guanine deaminase and guanosine kinase activities was found in neuroblastoma cells. Hypoxanthine was a source for both adenine and guanine nucleotides, whereas adenine or guanine were principally sources for adenine (>85%) or guanine (>90%) nucleotides, respectively. The rate of [¹⁴C]formate incorporation into ATP, GTP and nucleic acid purines was essentially equivalent for both N4 and N4TG cells. Purine nucleotide pools were also comparable in both cell lines, but the concentration of UDP-sugars was 1.5 times greater in N4TG than N4 cells.     

Metabolism of Guanine and Guanine Nucleotides in Primary Rat Neuronal Cultures

The metabolic fate of guanine and of guanine ribonucleotides (GuRNs) in cultured rat neurons was studied using labeled guanine. 8-Aminoguanosine (8-AGuo), an inhibitor of purine nucleoside phosphorylase, was used to clarify the pathways of GMP degradation, and mycophenolic acid, an inhibitor of IMP dehydrogenase, was used to assess the flux from IMP to GMP and, indirectly, the activity of the guanine nucleotide cycle (GMP----IMP----XMP----GMP). The main metabolic fate of guanine in the neurons was deamination to xanthine, but significant incorporation of guanine into GuRNs, at a rate of approximately 8.5-13.1% of that of the deamination, was also demonstrated. The turnover rate of GuRNs was fast (loss of 80% of the radioactivity of the prelabeled pool in 22 h), reflecting synthesis of nucleic acids (32.8% of the loss in radioactivity) and degradation to xanthine, guanine, hypoxanthine, guanosine, and inosine (49.3, 4.3, 4.1, 1.1, and 0.5% of the loss, respectively). Of the radioactivity in GuRNs, 7.9% was shifted to adenine nucleotides. The accumulation of label in xanthine indicates (in the absence of xanthine oxidase) that the main degradative pathway from GMP is that to xanthine through guanosine and guanine. The use of 8-AGuo confirmed this pathway but indicated the operation of an additional, relatively slower degradative pathway, that from GMP through IMP to inosine and hypoxanthine. Hypoxanthine was incorporated mainly into adenine nucleotide (91.5%), but a significant proportion (6%) was found in GuRNs.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of nutritional state and allopurinol on nucleotide formation in enterocytes from the guinea pig small intestine

The uptake of purine nucleosides (guanosine and hypoxanthine) and bases (guanine, hypoxanthine and adenine) and their incorporation into nucleotides were studied in enterocytes isolated from fed and 3-day fasted guinea pig jejunum. Both total uptake and synthesis of nucleotides were greater for these purines in the fasted, as compared to the fed state for the first 5 min, when the initial substrate concentration in the medium was 10 microM. Increased uptake did not result from a change in the relative distribution of synthesized nucleotides between the fed and fasted states. Reduced catabolism was observed in the medium by enterocytes from fasted as compared to fed animals after 1 min of incubation with both inosine and guanosine. Preincubation of enterocytes with allopurinol (a xanthine oxidase inhibitor) decreased total uptake but increased the formation of IMP from hypoxanthine. Xanthine oxidase activity measured in mucosa from fasted guinea pigs was lower than that from fed animals (6.29 vs. 9.30 nmol/min per mg protein, respectively). However, activities of the salvage enzymes adenine phosphoribosyltransferase and hypoxanthine-guanine phosphoribosyltransferase were not significantly different between the fed and fasted states. These data show that allopurinol treatment, and mucosal atrophy resulting from fasting, decrease xanthine oxidase activity and increase nucleotide synthesis from exogenous substrates in enterocytes from the guinea-pig small intestine, suggesting a regulatory function of mucosal xanthine oxidase in purine salvage by the small intestine.     

Purine requirements for the expression of Cape buffalo serum trypanocidal activity

Cape buffalo serum contains xanthine oxidase which generates trypanocidal H₂O₂ during the catabolism of hypoxanthine and xanthine. The present studies show that xanthine oxidase-dependent trypanocidal activity in Cape buffalo serum was also elicited by purine nucleotides, nucleosides, and bases even though xanthine oxidase did not catabolize those purines. The paradox was explained in part, by the presence in serum of purine nucleoside phosphorylase and adenosine deaminase, that, together with xanthine oxidase, catabolized adenosine, inosine, hypoxanthine, and xanthine to uric acid yielding trypanocidal H₂O₂. In addition, purine catabolism by trypanosomes provided substrates for serum xanthine oxidase and was implicated in the triggering of xanthine oxidase-dependent trypanocidal activity by purines that were not directly catabolized to uric acid in Cape buffalo serum, namely guanosine, guanine, adenine monophosphate, guanosine diphosphate, adenosine 3′:5-cyclic monophosphate, and 1-methylinosine. The concentrations of guanosine and guanine that elicited xanthine oxidase-dependent trypanocidal activity were 30–270-fold lower than those of other purines requiring trypanosome-processing which suggests differential processing by the parasites.     

Interconversion and uptake of nucleotides, nucleosides, and purine bases by the marine bacterium MB22.

Whole cells and isolated membranes of the marine bacterium MB22 converted nucleotides present in the external medium rapidly into nucleosides and then into bases. Nucleosides and purine bases formed were taken up by distinct transport systems. We found a high-affinity common transport system for adenine, guanine, and hypoxanthine, with a Km of 40 nM. This system was inhibited noncompetitively by purine nucleosides. In addition, two transport systems for nucleosides were present: one for guanosine with a Km of 0.8 microM and another one for inosine and adenosine with a Km of 1.4 microM. The nucleoside transport systems exhibited both mixed and noncompetitive inhibition by different nucleosides other than those translocated; purine and pyrimidine bases had no effect. The transport of nucleosides and purine bases was inhibited by dinitrophenol or azide, thus suggesting that transport is energy dependent. Inside the cell all of the substrates were converted mainly into guanosine, xanthine, and uric acid, but also anabolic products, such as nucleotides and nucleic acids, could be found.     

Purine metabolism by intracellular Chlamydia psittaci.

Purine metabolism was studied in the obligate intracellular bacterium Chlamydia psittaci AA Mp in the wild type and a variety of mutant host cell lines with well-defined deficiencies in purine metabolism. C. psittaci AA Mp cannot synthesize purines de novo, as assessed by its inability to incorporate exogenous glycine into nucleic acid purines. C. psittaci AA Mp can take ATP and GTP, but not dATP or dGTP, directly from the host cell. Exogenous hypoxanthine and inosine were not utilized by the parasite. In contrast, exogenous adenine, adenosine, and guanine were directly salvaged by C. psittaci AA Mp. Crude extract prepared from highly purified C. psittaci AA Mp reticulate bodies contained adenine and guanine but no hypoxanthine phosphoribosyltransferase activity. Adenosine kinase activity was detected, but guanosine kinase activity was not. There was no competition for incorporation into nucleic acid between adenine and guanine, and high-performance liquid chromatography profiles of radiolabelled nucleic acid nucleobases indicated that adenine, adenosine, and deoxyadenosine were incorporated only into adenine and that guanine, guanosine, and deoxyguanosine were incorporated only into guanine. Thus, there is no interconversion of nucleotides. Deoxyadenosine and deoxyguanosine were cleaved to adenine and guanine before being utilized, and purine (deoxy)nucleoside phosphorylase activity was present in reticulate body extract.     

Purine metabolite concentrations in portal and peripheral blood of steers, sheep and rats.

1. The concentration of purine derivatives in portal and peripheral blood of steers, sheep and rats was measured by reverse-phase high performance liquid chromatography. 2. Nucleotides, nucleosides (apart from inosine), adenine and guanine were not found in the plasma samples. Allantoin, uric acid, hypoxanthine and xanthine accounted for virtually all purine metabolites in plasma samples. 3. Non-oxidized derivatives (hypoxanthine and xanthine) were consistently detected in sheep but not in steer or rat plasma samples showing a differential availability of reutilizable purine derivatives between species.     

Purine metabolism and urate biosynthesis in isolated chicken hepatocytes

The metabolism of some purine compounds to urate and their effects on de novo urate synthesis in chicken hepatocytes were investigated. The purines, listed in descending order of rates of catabolism to urate, were hypoxanthine, xanthine, inosine, guanosine, guanine, IMP, GMP, adenosine, AMP, and adenine. During a 1-h incubation period, conversion to urate accounted for more than 80% of the total quantities of guanine, guanosine, and inosine metabolized, but only 42% of the adenosine and 23% of the adenine metabolism. Adenine, adenosine, and AMP inhibited de novo urate synthesis [( 14C]formate incorporation into urate), whereas the other purines, especially guanine, guanosine, and GMP, stimulated de novo urate synthesis. When hepatocytes were incubated with glutamine and adenosine, AMP, guanine, guanosine, or GMP, the rates of de novo urate synthesis were lower than the additive effects of glutamine and the purine in separate incubations. Increasing phosphate concentrations had no effect on urate synthesis in the absence of added purines but, in combination with adenosine, AMP, guanosine, or GMP, increased urate synthesis. These results indicate that the ratio of adenine to guanine nucleotides and the interaction between substrates and purine nucleotides are involved in the regulation of urate biosynthesis in chicken liver.     

Biochemistry and metabolism of Giardia

Giardia lamblia, an aerotolerant anaerobe, respires in the presence of oxygen by a flavin, iron-sulfur protein-mediated electron transport system. Glucose appears to be the only sugar catabolized by the Embden-Meyerhof-Parnas and hexose monophosphate pathways, and energy is produced by substrate level phosphorylation. Substrates are incompletely oxidized to CO2, ethanol and acetate by nonsedimentable enzymes. The lack of incorporation of inosine, hypoxanthine, xanthine, formate or glycine into nucleotides indicates an absence of de novo purine synthesis. Only adenine, adenosine, guanine and guanosine are salvaged, and no interconversion of these purines was detected. Salvage of these purines and their nucleosides is accomplished by adenine phosphoribosyltransferase, adenosine hydrolase, guanosine phosphoribosyltransferase and guanine hydrolase. The absence of de novo pyrimidine synthesis was confirmed by the lack of incorporation of bicarbonate, orotate and aspartate into nucleotides, and by the lack of detectable levels of the enzymes of de novo pyrimidine synthesis. Salvage appears to be accomplished by the action of uracil phosphoribosyltransferase, uridine hydrolase, uridine phosphotransferase, cytidine deaminase, cytidine hydrolase, cytosine phosphoribosyltransferase and thymidine phosphotransferase. Nucleotides of uracil may be converted to nucleotides of cytosine by cytidine triphosphate synthetase, but thymidylate synthetase and dihydrofolate reductase activities were not detected. Uptake of pyrmidine nucleosides, and perhaps pyrimidines, appears to be accomplished by carrier-mediated transport, and the common site for uptake of uridine and cytidine is distinct from the site for thymidine. Thymine does not appear to be incorporated into nucleotide pools. Giardia trophozoites appear to rely on preformed lipids rather than synthesizing them de novo.(ABSTRACT TRUNCATED AT 250 WORDS)     

Preservation of red blood cells with purines and nucleosides. II. Uptake and utilization of purines and nucleosides by stored red blood cells

The uptake of adenine, guanine, guanosine and inosine by stored red cells was investigated in whole blood and red cell resuspensions at initial concentrations of 0.25, 0.5 and 0.75 mM for adenine and 0.5 mM for the other additives using a rapid ion-exchange chromatographic microanalysis of purines and nucleosides in plasma and whole blood. Increasing adenine concentrations from 0.25 to 0.75 mM in blood elevated the adenine uptake from 0.3 up to 0.8 mmol/l red cells during 2 hours after collecting blood. The intra-/extracellular distribution ratio changed from 1 : 1.3 to 1: 1.7. Some 2 hours after withdrawing blood into CPD--solution with purines and nucleosides the uptake of adenine and guanine resulted in 40 per cent and 70 per cent respectively and of guanosine and inosine in 80 and 90 per cent respectively. The replacement of plasma by a resuspending solution gave the same uptake rates for purines and nucleosides. The nucleosides were rapidly split to purines and R-1-P and disappeared from blood during one week. Adenine and guanine were utilized to 80 to 90 per cent only after 3 weeks. During the same period the utilization of guanine was smaller by 40 per cent than that of adenine due to the different activity of the purine nucleoside phosphorylase for these substrates. The plasma of all analyzed blood samples contained hypoxanthine and inosine, but guanine and guanosine were detected only in those samples to which one of them was added. After 3 weeks of storage the highest concentration of hypoxanthine was found in CPD-AI blood with 600 microM in plasma and the highest concentration of synthesized inosine in CPD-AG blood with a concentration of 100 microM in plasma. Three ways of utilization of purines by stored red cells were discussed : the synthesis of nucleotide monophosphates, the formation of nucleosides, and the deamination. The portions of these ways change during storage. The most effective concentrations of adenine and guanosine in stored blood seems to be 0.25 and 0.5 mM respectively. The full utilization of the nucleoside requires the addition of inorganic phosphate.     

Entamoeba histolytica: uptake of purine bases and nucleosides during axenic growth.

A comparison was made of the uptake mechanisms of selected purine bases and nucleosides by axenically grown Entamoeba histolytica. Adenine, adenosine, and guanosine were taken up, in part, by a “carrier”-mediated system. Guanine, hypoxanthine, and inosine entered amoebas via diffusion. Inhibitor studies support the presence of individual transport sites for adenine-adenosine and adenosine-guanosine. Additional sites for transport of adenine, adenosine, and guanosine are implied by “non-productive binding” involving guanine, hypoxanthine, and inosine. Uptake of adenine, adenosine, and guanosine was reduced by iodoacetate and N-ethylmaleimide. Ribose failed to inhibit uptake of purine nucleosides.     

Sugar-free growth of mammalian cells on some ribonucleosides but not on others

It was shown earlier that a variety of vertebrate cells could grow indefinitely in sugar-free medium supplemented with either uridine or cytidine at greater than or equal to 1 mM. In contrast, most purine nucleosides do not support sugar-free growth for one of the following reasons. The generation of ribose-1-P from nucleoside phosphorylase activity is necessary to provide all essential functions of sugar metabolism. Some nucleosides, e.g. xanthosine, did not support growth because they are poor substrates for this enzyme. De novo pyrimidine synthesis was inhibited greater than 80% by adenosine or high concentrations of inosine, e.g. 10 mM, which prevented growth on these nucleosides; in contrast, pyrimidine synthesis was inhibited only marginally on 1 mM inosine or guanosine, but normal growth was only seen on 1 mM inosine, not on guanosine. The inhibition of de novo adenine nucleotide synthesis prevented growth on guanosine, since guanine nucleotides could not be converted to adenine nucleotides. Guanine nucleotides were necessary for this inhibition of purine synthesis, since a mutant blocked in their synthesis grew normally on guanosine. De novo purine synthesis was severely inhibited by adenosine, inosine, or guanosine, but in contrast to guanosine, adenosine and inosine could provide all purine requirements by direct nucleotide conversions.     

Salvage synthesis of purine nucleotides by Helicobacter pylori

G.L. MENDZ, B.M. JIMENEZ, S.L. HAZELL, A.M. GERO AND W.J. O'SULLIVAN. 1994. The incorporation of purine nucleotide precursors into Helicobacter pylori and the activities of enzymes involved in nucleotide salvage biosynthetic pathways were investigated by radioactive tracer analysis and nuclear magnetic resonance spectroscopy. The organism took up the nucleobases adenine, guanine and hypoxanthine, and the nucleosides adenosine, guanosine and deoxyadenosine. Any incorporation of deoxyguanosine by the cells was below the detection limits of the methods employed. The activities of adenine-, guanine- and hypoxanthine-phosphoribosyl transferases were established. The bacterium showed high levels of adenosine and guanosine nucleosidase activities and lesser activity for deoxyadenosine; no hydrolysis of deoxyguanosine was detected. Phosphorylase activities were not observed with any of the nucleosides. Phosphotransferase activities with similar rates were demonstrated for adenosine, guanosine and deoxyadenosine; and a weaker activity was detected for deoxyguanosine. No nucleoside kinase activities were observed with any of the nucleosides. The presence of adenylate kinase was established, but no guanylate kinase activity was observed. The study provided evidence for the presence in H. pylori of salvage pathways for the biosynthesis of purine nucleotides.