Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Formycins”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 289 records · Page 16Linked to original sources
PubMed1994

Expression of a renal Na(+)-nucleoside cotransport system (N2) in Xenopus laevis oocytes.

Xenopus laevis oocytes have been used for the expression of a renal, pyrimidine-selective, Na(+)-nucleoside cotransporter (N2). As compared to its uptake in water-injected oocytes, Na(+)-dependent thymidine uptake was enhanced in a time- and dose-dependent manner in oocytes injected with rat renal cortex total poly(A)+ RNA. An increased uptake was also observed after injection of size fractionated rat renal cortex poly(A)+ RNA (2-3 kb). Consistent with the selectivity of the N2 nucleoside transporter, cytidine significantly inhibited Na(+)-dependent thymidine uptake in oocytes injected with total poly(A)+ RNA whereas guanosine and formycin B did not. Na(+)-dependent thymidine uptake was also enhanced in oocytes injected with size fractionated human renal cortex poly(A)+ RNA (2-3 kb). The above data demonstrate functional expression of renal cortex, Na(+)-nucleoside cotransporters in Xenopus laevis oocytes.

Animals↗
PubMed1980

Adenosine deaminase from Azotobacter vinelandii. Purification and properties.

Adenosine deaminase (EC 3.5.4.4) was found to occur in the extract of Azotobacter vinelandii, strain 0, and purified by heating at 65 degrees C, fractionation with ammonium sulfate, DEAE-cellulose chromatography and gel filtration on Sephadex G-150. Purified adenosine deaminase was effectively stabilized by the addition of ethylene glycol. The molecular weight of the enzyme was estimated to be 66,000 by gel filtration on Sephadex G-150. The enzyme specifically attacked adenosine and 2'-deoxyadenosine to the same extent, and formycin A to a lesser extent. The pH optimum of the enzyme was observed at pH 7.2. Double reciprocal plot of initial velocity versus adenosine concentration was concave upward, and Hill interaction coefficient was calculated to be 1.5, suggesting the allosteric binding of the substrate. ATP inhibited adenosine deaminase in an allosteric manner, whereas other nucleotides were without effect. The physiological significance of the enzyme was discussed in relation to salvage pathway of purine nucleotides.

Adenosine Deaminase↗
PubMed1987

Inosine partially mimics the effects of glucose on ionic fluxes, electrical activity, and insulin release in mouse pancreatic B-cells.

The purine ribonucleoside inosine is known to be metabolized in islet cells (its ribose moiety feeds into the pentose-phosphate cycle) and stimulate insulin release, but the mechanisms of this stimulation have not been established. These were investigated with mouse islets. In the absence of glucose, 5 mM inosine decreased 86Rb+ efflux from islet cells, depolarized the B-cell membrane, induced electrical activity (slow waves of membrane potential with bursts of spikes on the plateau), accelerated 45Ca2+ efflux and stimulated insulin release with the same efficiency as 10 mM glucose. Raising the concentration of inosine to 20 mM only had a slight further effect and, in particular, failed to cause persistent depolarization of the B-cell membrane. The electrical activity triggered by inosine was blocked by cobalt, and the stimulation of 45Ca2+ efflux and insulin release was abolished in a Ca2+-free medium. The effects of 10 mM glucose on electrical activity, 45Ca2+ efflux and insulin release were augmented by as little as 0.5 mM inosine. All effects of inosine were abolished by an inhibitor of nucleoside transport (nitrobenzylthioguanosine) and markedly impaired by inhibitors of nucleoside phosphorylase (formycin B) or of glycolysis (iodoacetate). In conclusion, inosine metabolism in B-cells induces insulin release by triggering the same sequence of events as glucose metabolism: a decrease of K+ permeability of the B-cell membrane, leading to depolarization and activation of voltage-dependent Ca channels.

Animals↗
PubMed1976

Poly ADP-ribosylation of DNA-dependent RNA polymerase I from quail oviduct. Dependence on progesterone stimulation.

Progesterone causes in goblet cells of oviducts of estrogen hormone-stimulated immature quails selectively gene activation without affecting DNA synthesis. This biological model has been used to study the influence of poly ADP-ribosylation during the processes of DNA transcription. Administration of progesterone in vivo causes an increase of the activity of RNA polymerase I and II in isolated nuclei. This increase is accompanied by a marked decrease of the specific activity of poly (ADP-Rib) polymerase. After in vitro ADP-ribosylation of nuclear proteins the template capacity of chromatin for ""exogenous'' RNA synthesis (with E. coli DNA-dependent RNA polymerases) as well as for ""endogenous'' RNA synthesis with DNA dependent RNA polymerases II is not affected, whereas the data presented seem to indicate that the capacity for RNA synthesis mediated by ""endogenous'' DNA-dependent RNA polymerase I might be inhibited after ADP-ribosylation. Evidence is presented to show that a considerable amount of poly (ADP-Rib), synthesized by poly (ADP-Rib) polymerase in isolated nuclei, is linked with RNA polymerase I. The rate of synthesis of poly (ADP-Rib) is dependent on the incubation temperature (optimum at 25 degrees C) and it can be inhibited by the specific inhibitors of poly (ADP-Rib) polymerase nicotineamide, thymidine and formycin B. Poly (ADP-Rib) is probably associated with RNA polymerase I through a covalent linkage. ADP-ribosylated RNA polymerase I has been purified 550 fold with respect to the nuclear extract corresponding to a 4,000 fold purification from the whole cell homogenate. The ratio between poly (ADP-Rib), formed during preincubation of nuclei with NAD, and RNA polymerase I remains almost constant during the purification procedures. The extent of ADP-ribosylation of RNA polymerase I decreases during gene expression. Thus we conclude that poly ADP-ribosylation of this enzyme is one of the regulatory mechanisms by which specificity of DNA transcription is achieved.

Animals↗
PubMed2004

Uptake of the anthracycline pirarubicin into mouse M5076 ovarian sarcoma cells via a sodium-dependent nucleoside transport system.

PURPOSE: We have previously demonstrated that the cytotoxicity of anthracyclines, pirarubicin (THP) and doxorubicin (DOX), is partially dominated by their intracellular amounts, which depend on the uptake efficacy of transporter(s). To clarify their transport mechanism, we examined whether or not Na+/nucleoside cotransporter (CNT) is involved in the uptake of THP by M5076 cells. METHODS: Expression of the CNT isoforms was determined by reverse-transcription PCR. We used two cell lines, intact M5076 and CNT2-transfected Cos-7 cells, to characterize the uptake of THP and [3H]uridine. RESULTS: The mRNA for CNT2, but not that for CNT1 or CNT3, was expressed in M5076 cells, and [3H]uridine uptake by the cells required a Na+ gradient as a driving force. THP uptake by M5076 cells depended on a Na+ gradient, and furthermore, formycin B and AZT had cis-inhibitory and trans-stimulatory effects on the uptake. The efflux of [3H]uridine from M5076 cells was stimulated by the addition of THP extracellularly, which constituted definite evidence of CNT-mediated uptake of THP. However, THP uptake by CNT2 transfectant was almost the same as that by mock cells, indicating that an unidentified CNT isoform contributes to THP uptake by M5076 cells, this being supported by the differences in transport characteristics of [3H]uridine between M5076 and CNT2-transfected cells. CONCLUSION: THP is partially taken up into M5076 cells via a novel Na+-dependent transport system common to nucleosides.

Animals↗
PubMed1984

Adenylosuccinate synthetase from Dictyostelium discoideum: effects of hadacidin analogs and binding of [14C]hadacidin.

The enzyme adenylosuccinate (sAMP) synthetase has been partially purified from Dictyostelium discoideum using hadacidin-Sepharose 4B affinity chromatography, anion-exchange high-performance liquid chromatography (HPLC), and gel-filtration HPLC, resulting in a 2600-fold purification. Using a newly developed HPLC procedure to assay activity, it has been found that D. discoideum adenylosuccinate synthetase activity has apparent Km values for the substrates IMP, GTP, and aspartate of 36, 23, and 714 microM, respectively. The analog guanosine-5'-(beta, gamma-imino)triphosphate was found to be an inhibitor of GTP with a Ki of 15 microM, and IMP was competitively inhibited by its analog formycin B monophosphate with a Ki of 80 microM. An analysis of these kinetic data showed a pattern consistent with a fully random terter mechanism. Hadacidin, an analog of aspartate, was an inhibitor of that substrate at 86 microM. Other analogs of hadacidin were synthesized and examined for their effect on the sAMP synthetase activity. Compared to hadacidin, which produced 100% inhibition at 5 mM, it was observed that N-acetyl-N-hydroxyglycine, N-formylglycine, N-acetylglycine, and N-hydroxyglycine all inhibited between 50 and 75%, with N-(thiocarboxy)-L-aspartic anhydride less effective at 27%, and N-benzoylglycine at only 6%. N-Formylsarcosine, N-acetylmethionine, O-methylpyruvate oxime, and hadacidin methylester had no effect at this concentration. The adenylosuccinate synthetase activity was dependent on metal ions with maximum activity being obtained with Mg2+. The ability of the aspartate analog hadacidin to bind to the purified adenylosuccinate synthetase was demonstrated using anion-exchange HPLC and [formyl-14C]hadacidin. The radioactivity coeluted with the adenylosuccinate synthetase and the bound, radiolabeled hadacidin was displaced by excess aspartate.

Adenylosuccinate Synthase↗
PubMed1978

Specificity of nucleotide binding and coupled reactions utilising the mitochondrial ATPase.

1. Tightly bound ATP and ADP, found on the isolated mitochondrial ATPase, exchange only slowly at pH 8, but the exchange is increased as the pH is reduced. At pH 5.5, more than 60% of the bound nucleotide exchanges within 2.5 min. 2. Preincubation of the isolated ATPase with ADP leads to about 50% inhibition of ATP hydrolysis when the enzyme is subsequently assayed in the absence of free ADP. This effect, which is reversed by preincubation with ATP, is absent on the membrane-bound ATPase. This inhibition seems to involve the replacement of tightly bound ATP by ADP. 3. Using these two findings, the binding specificity of the tight nucleotide binding sites was determined. iso-Guanosine, 2'-deoxyadenosine and formycin nucleotides displaced ATP from the tight binding sites, while all other nucleotides tested did not. The specificities of the tight sites of the isolated and membrane-bound ATPase were similar, and higher than that of the hydrolytic site. 4. The nucleotide specificities of 'coupled processes' nucleoside triphosphate-driven reversal of electron transfer, nucleoside triphosphate-32Pi exchange and phosphorylation were higher than that of the hydrolytic site of the ATPase and similar to that of the tight nucleotide binding sites.

Adenosine Diphosphate↗
PubMed1990

Mobility of nucleoside transporter of human erythrocytes differs greatly when loaded with different nucleosides.

Time courses of transmembrane equilibration of 2-chloroadenosine, 2'-deoxyadenosine, 3'-deoxyadenosine, cytidine and 2'-deoxycytidine were measured by rapid kinetic techniques in human erythrocytes under equilibrium exchange and zero-trans conditions. The kinetic parameters for transport were computed by fitting appropriate integrated rate equations to the data pooled for seven concentrations and compared to the kinetic parameters for uridine, adenosine, thymidine and formycin B transport determined previously for human erythrocytes under comparable experimental conditions. The transport of all nucleosides conformed to the simple carrier model and was directionally symmetric. The Michaelis-Menten constants for equilibrium exchange (Kee) ranged from 22 microM for 2-chloroadenosine to about 4 mM for cytidine and the maximum velocities (Vee) differed in a similar manner, so that the first-order rate constants (Vee/Kee) were similar for all nucleosides. The kinetic parameters for 2'-deoxyadenosine transport were similar to those for adenosine transport, whereas the lack of the 3'-OH group greatly reduced the affinity of 3'-deoxyadenosine (cordycepin) for the carrier. 2', 3'-Dideoxynucleosides were transported less than 1% as efficiently as 2'- and 3'-deoxynucleosides. Thus, the 2'- and 3'-OH groups play an important role in nucleoside transport. The mobility of the carrier when loaded with pyrimidine nucleosides (reflected by Vee) was 5-10-times greater than that of the empty carrier, whereas the mobility of the adenosine-loaded or 2'-deoxyadenosine-loaded carrier was about equal to that of the empty carrier. Loading the carrier with 2-chloroadenosine or 3'-deoxyadenosine actually decreased its mobility. Thus, the differential mobility of the loaded and empty carrier differs greatly with the nucleoside substrate. The mobility of the loaded carrier as well as Kee increased with a decrease in lipid solubility of the nucleoside substrate, but the relationship was complex.

2-Chloroadenosine↗
PubMed1993

Substrate selectivity, potential sensitivity and stoichiometry of Na(+)-nucleoside transport in brush border membrane vesicles from human kidney.

Recently, we demonstrated the presence of a Na(+)-nucleoside cotransport mechanism that transports both purine and pyrimidine nucleosides in human renal brush-border membrane vesicles (BBMV) (Gutierrez et al. (1992) Biochim. Biophys. Acta 1105, 1-9). The objective of this study was to further elucidate the characteristics of this cotransport system in terms of electrical potential sensitivity, stoichiometry and substrate selectivity with respect to nucleoside analogs. In BBMV from human kidney, Na(+)-thymidine uptake was stimulated by an inside negative potential difference created by K+ and valinomycin. A hyperbolic relationship between initial rate of uridine uptake and Na+ concentration was obtained suggesting a Na(+)-nucleoside coupling stoichiometry of 1:1. Our previous study had demonstrated that the pyrimidines, thymidine, cytidine, and uridine and the purines, adenosine, 2'-deoxyadenosine, and guanosine, but not inosine and formycin B, were substrates of this system. To further define the substrate selectivity of the transporter, the interaction of the drugs, 2-chloroadenosine (2-ClAdo), 5-fluorouridine (5-FUrd) and 5-iodo-2'-deoxyuridine (5-IdUrd), nucleoside analogs that are modified on the base moiety was studied. The three compounds inhibited Na(+)-thymidine uptake in the vesicles via a competitive mechanism. The IC50 values for 2-ClAdo, 5-FUrd and 5-IdUrd were 75, 49, and 16 microM, respectively. In addition, 5-IdUrd trans-stimulated the initial uptake of thymidine into the vesicles suggesting that the two compounds share the same transporter. Collectively, these data suggest that Na(+)-nucleoside transport in the human renal brush-border membrane is an electrogenic process and that the kidney may play a role in the disposition and targeting of clinically important nucleoside analogs.

2-Chloroadenosine↗
PubMed1978

Characterization and comparison of poly(adenosine dephosphoribose) synthesis and DNA synthesis in nucleotide-permeable cells.

Using eukaryotic cells that have been rendered permeable to exogenously supplied nucleotides, we have characterized the activity of the poly(adenosine diphosphoribose) (poly(ADPR)) synthesis system and compared it to the DNA synthesis complex. The synthesis of poly(ADPR) is dependent on the presence of NAD and Mg2+. It does not require ATP, NaF or a monovalent cation. It is inhibited by N-ethylmaleimide. The reaction product conforms to the nuclease susceptibilities expected for poly(ADP ribose) in that it is degraded by venom phosphodiesterase but not by DNAase of RNAase. A comparison of the effects of inhibitors of poly(ADPR) synthesis and DNA synthesis clearly distinguishes between the two enzymatic systems. Nicotinamide, 5-methyl nicotinamide, thymidine, 5-bromo deoxyuridine, adenosine diphosphoribose, caffeine and formycin all inhibit poly(ADPR) synthesis but not DNA synthesis. In contrast, araCTP, cytembena and phosphonoacetic acid all inhibit DNA synthesis but not poly(ADPR) synthesis. Addition of DNAase to the permeable cells causes a marked stimulation of poly(ADPR) synthesis. L cells in logarithmic growth were found to have high levels of activity of the DNA synthesis complex and low levels of activity of the poly(ADPR) synthesis system. In contrast, cells at plateau phase density demonstrate a decrease in the activity of the DNA synthesis complex and a marked increase in activity of the poly(ADPR) synthesis system. When examined in the presence of added DNAase, the activity of the poly(ADPR) synthesis system is the same in cells obtained from log or plateau phase cultures. This indicates that the physiologic activity of the enzyme varies while the total amount of enzyme remains constant. When the permeable cells are allowed to synthesize both poly(ADPR) and DNA simultaneously, the synthesis of one polymer has no effect on the rate of synthesis of the other.

Cell Cycle↗
PubMed1985

Differential incorporation of 2'-deoxyadenosine into human peripheral lymphocytes.

Human peripheral lymphocytes incubated with 2'-deoxycoformycin and 2'-deoxyadenosine (dAdo) reached a plateau in dATP accumulation after 4 hr that lasted for up to 24 hr. Total dATP accumulation did not exceed 15% of the control ATP concentration in the lymphocytes. In contrast, the human CCRF-CEM T lymphoblastic cell line and human erythrocytes showed a nearly linear pattern of dATP formation throughout the incubation period. By 6 hr the dATP concentration in the CCRF-CEM cells exceeded the control ATP concentration. A comparison of dATP accumulation in purified peripheral T and B lymphocytes indicated differences between these cells that favor greater dATP formation in the B lymphocytes. Incorporation studies with several adenosine analogs demonstrated that arabinosyladenine, 2-F-arabinosyladenine, tubercidin, formycin A, and 9-(2'-deoxy-2'-fluoro-beta-D-ribofuranosyl)adenine form corresponding amounts of analog triphosphate in the T and B cell-enriched lymphocytes. 9-(2'-Deoxy-2'-fluoro-beta-D-arabinofuranosyl)adenine (2'-F-araA) was the only compound to show an incorporation pattern similar to that observed with dAdo by forming analog triphosphate only in the B cell-enriched lymphocyte population. Nucleoside kinase measurements showed no significant differences in dAdo, adenosine, or 2'-deoxycytidine kinase activities between the T and B lymphocytes. The inability of the T cells to incorporate dAdo or the analog 2'-F-araA into their nucleotide pools may indicate the existence of a highly specific catabolic enzyme(s).

Adenosine Deaminase↗
PubMed1990

Inhibition of nucleoside transport by nitrobenzylthioformycin analogs.

The formycin analogs of nitrobenzylthioinosine and nitrobenzylthioguanosine were synthesized and evaluated as nucleoside transport inhibitors. These analogs have a potential therapeutic advantage over their parent compounds in that their C-nucleosidic linkages prevent them from being degraded to the immunosuppressive agents, 6-mercaptopurine and 6-thioguanine. 7-[(4-Nitrobenzyl)-thio]-3-(beta-D-ribofuranosyl)pyrazolo[4,3- d]pyrimidine (NBTF) and 5-amino-7-[(4-nitrobenzyl)thio]-3-(beta-D- ribofuranosyl)pyrazolo[4,3-d]pyrimidine (NBTGF) were inhibitors of nucleoside transport in human erythrocytes and HL-60 leukemia cells. The IC50 value for nitrobenzylthioinosine, NBTF and NBTGF with 10% erythrocyte suspensions were 18, 18 and 40 nM respectively. Specific binding studies with [3H]NBTF yielded a Kd of 3.4 nM with erythrocytes, approximately 10-fold higher than values reported for nitrobenzylthioinosine. NBTF and nitrobenzylthioinosine bound to HL-60 cells with Kd values of 8.1 and 0.81 nM respectively. The octanol/water partition coefficients of nitrobenzylthioinosine, NBTF and NBTGF were 3.5, 3.2, and 2.8 respectively. NBTF could be expected to be equipotent with nitrobenzylthioinosine in whole blood where inhibitor concentrations of 10(-7) to 10(-6) M are required in order to saturate erythrocytic binding sites; hence, it may exhibit the advantages inherent in a C-nucleoside.

Adenosine↗
PubMed1994

Expression of a human renal sodium nucleoside cotransporter in Xenopus laevis oocytes.

In this study, Xenopus laevis oocytes injected with poly(A)+ RNA (mRNA) isolated from human kidney were used to express a Na(+)-nucleoside cotransporter. Na(+)-stimulated [3H]thymidine uptake was enhanced 2-3-fold in oocytes injected with 50 ng poly(A)+ RNA and 4-5-fold in oocytes injected with 20 ng of a size-fractionated human renal cortex mRNA fragment (2-3 kb) in comparison with water-injected oocytes. Na(+)-dependent thymidine uptake in oocytes injected with the 2-3 kb mRNA fragment was inhibited significantly by thymidine and guanosine but not by formycin B, consistent with the N4 Na(+)-nucleoside cotransporter. The Km (28 microM) of Na(+)-dependent thymidine uptake in the oocytes injected with the 2-3 kb mRNA fragment was similar to the Km (27 microM) of Na(+)-dependent thymidine uptake obtained in human renal brush border membrane vesicles. These data suggest for the first time that a Na(+)-nucleoside cotransporter from human kidney can be expressed in X. laevis oocytes.

Animals↗
PubMed1989

Leishmania donovani: isolation and characterization of sodium stibogluconate (Pentostam)-resistant cell lines.

Leishmania donovani promastigotes were generated by virtue of their resistance to incrementing concentrations of sodium stibogluconate (Pentostam) under completely defined growth conditions. The PENT0400 and PENT03200 cell lines were isolated after prolonged exposure to 0.4 mg/ml and 3.2 mg/ml Pentostam (Sb concentration), respectively. Whereas the effective concentration of Pentostam which inhibited the growth of wild type cells by 50% (EC50 value) was 0.1-0.15 mg/ml, the EC50 values for the PENT0400 and PENT03200 cells were approximately 1 and 4 mg/ml, respectively. The decreased sensitivities of both PENT0400 and PENT03200 cells to Pentostam were maintained after 6 months of continuous culture in the absence of selective pressure, indicating that the Pentostam resistance in the mutant organisms was a stable genetic trait. Interestingly, wild type and PENT03200 cells were equally sensitive to growth inhibition and cytotoxicity caused by SbCl5 and SbCl3, as well as to a variety of other cations such as Cd, Zn, and As. Wild type and PENT03200 cells also displayed equivalent growth sensitivities to a spectrum of other antiprotozoal agents, including antimony potassium tartrate, melarsoprol, pyrimethamine, pentamidine, formycin B, and difluoromethylornithine. These results illustrate a potentially useful model system to study Pentostam resistance in Leishmania and suggest that Pentostam resistance in vitro may be independent of antimony toxicity.

Animals↗
PubMed1991

Leishmania donovani: characteristics of adenosine and inosine transporters in promastigotes of two different strains.

The nucleoside transport characteristics of two strains of Leishmania donovani promastigotes were studied. Strain S1, growing in fully defined medium, and strain S2 (MHOM/ET/67/HA3) both transported adenosine and inosine, but only strain S1 transported uridine and thymidine. Competition studies in the presence of 100 microM of unlabeled adenosine, inosine, guanosine, 2'-deoxyadenosine, tubercidin, formycin B, 3'-deoxyinosine as well as uridine, thymidine and cytidine, with either 1 microM [3H]adenosine or [3H]inosine as permeant, were carried out. The inhibition profile with [3H]inosine as permeant was essentially identical in S1 and S2 promastigotes, indicating that the same inosine transporter was present in both strains. However, with [3H] adenosine as permeant, significant differences were noted between the two strains. Thus, only adenosine, 2'-deoxyadenosine, tubercidin, uridine, and thymidine were strongly inhibitory in S1 promastigotes, while essentially all nucleosides tested were effective in S2 promastigotes. This indicates that adenosine transport in S2 promastigotes seems to involve a transporter differing from that described for S1 promastigotes.

Adenosine↗
PubMed1995

5'-AMP inhibits dephosphorylation, as well as promoting phosphorylation, of the AMP-activated protein kinase. Studies using bacterially expressed human protein phosphatase-2C alpha and native bovine protein phosphatase-2AC.

Human protein phosphatase-2C alpha (PP2C alpha) was purified to homogeneity after expression in Escherichia coli. AMP inhibited the dephosphorylation of AMP-activated protein kinase (AMPK), but not phosphocasein, by PP2C alpha. The concentration dependence and the effects of other nucleotides (ATP and formycin A-5'-monophosphate) suggest that AMP acts by binding to the same site which causes direct allosteric activation of AMPK. A similar, although less pronounced, effect was observed with another protein phosphatase (PP2AC). We have now shown that AMPK activates the AMPK cascade by four mechanisms, which should make the system exquisitely sensitive to changes in AMP concentration.

AMP-Activated Protein Kinases↗
PubMed1991

Isolation and characterization of the protein phosphoamidates formed by a membrane bound adenylyl transferase reaction in Dictyostelium discoideum.

1. In this paper we report the results of studies on an adenylyl transferase (AyTase) activity from Dictyostelium discoideum. 2. Previous studies suggested that this activity catalyzed the transfer of AMP from ATP to a membrane protein to form a phosphoamidate reaction product. 3. In the present study we have isolated and characterized the product of the AyTase reaction and surprisingly found two AMP labeled products by SDS-gel-filtration HPLC. 4. The apparent molecular weights of these phosphoamidates were 13.5 and 1.5 kDa. 5. In addition, because the reaction product was rapidly degraded by a phosphoamidase, experiments were undertaken using AVAMP, a synthetic phosphoamidate, as an alternative phosphoamidase substrate, to trap the reaction products. 6. As the phosphoamidase activity could be inhibited by cAMP, cyclic formycin monophosphate, an analog of cAMP resistant to hydrolysis by cAMP phosphodiesterase, was also used to trap the products. 7. Both attempts at trapping failed. 8. A model for the AyTase reaction was developed to account for the failure to trap the products and the formation of two phosphoamidates.

Adenosine Monophosphate↗
PubMed1982

The purine metabolism of protozoa.

Protozoa possess a wealth of purine-salvage enzymes, many with unique, or unusual, substrate specificities. As a result, many opportunities for the chemotherapist exist. An exemplification is found in the conversion in schistosomes of allopurinol ribonucleoside to the corresponding ribonucleotide followed by further anabolism to the very toxic 4-aminopyrazolo(3,4-d)pyrimidine 1-ribonucleotide. The same organisms convert another inosine analog, formycin B, to the ribonucleotide, but its inhibitory effects appear to be exercised primarily by inhibition of the organism's adenylosuccinate synthase. A substantial segment of the Phylum Protozoa shows no vestigial traces of ability to synthesize purines de novo although thymidylate synthase appears to be present in many. The absence of other tetrahydrofolate catalyzed reactions suggests that these functions were never acquired.

Allopurinol↗