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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)

Adenine phosphoribosyltransferase has been purified to apparent homogeneity from mouse mammary tumor FM3A cells. The purified enzyme, with a specific activity of 20.6 X 10(6) units/g protein at 30 degrees C, was homogeneous as judged by polyacrylamide gel electrophoresis and Ouchterlony double immunodiffusion analysis. The native enzyme had a molecular weight of 44,000 and a subunit composition of 23,000. Apparent Km values for adenine and 5-phosphoribosyl-1-pyrophosphate (PRib-PP) were 6.6 microM and 1.2 microM, respectively. Free Mg2+ was an essential activator with a half-maximal effect at 0.4 mM. AMP was an inhibitor, competitive with PRib-PP, and the Ki value was estimated to be 24 microM. The enzyme activity was not significantly affected by 2,6-diaminopurine, 4-carbamoylimidazolium 5-olate, 8-azaadenine, and 2-fluoro-6-aminopurine. An antibody against the purified mouse adenine phosphoribosyltransferase was raised in a rabbit. The enzyme derived from either mouse, Chinese hamster, or human cells was completely neutralized and precipitated by this antibody, indicating that these enzymes share a common antigenic determinant.
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PMID:Purification and characterization of adenine phosphoribosyltransferase from mouse mammary carcinoma FM3A cells in culture. 376 21

Sealed and unsealed plasma membrane vesicles were prepared from human erythrocytes and lymphocytes. Phosphoribosylpyrophosphate synthetase (PRibPP synthetase), hypoxanthine phosphoribosyltransferase (HPRTase), and adenine phosphoribosyltransferase (APRTase) activities are detectable on both inside-out and right-side-out sealed vesicles. Ghost preparations were about 0.2%, 1%, and 1.2% of the total erythrocyte and 0.5%, 5.3%, and 9.7% of the lymphocyte APRTase, HPRTase, and PRibPP synthetase activities. The rapid decrease in these enzyme activities, upon further purification of the membranes, seemed to suggest that they might be loosely bound extrinsic proteins. Evidence confirming the localization of these enzymes on the cell surface was obtained by measuring production of [14C]AMP by intact cells in medium containing [14C]adenine, ribose 5-phosphate, and Mg2+ATP. The formation of AMP was linear with time and number of cells present. Magnesium and phosphate exerted different effects on the production of extracellular AMP than on intracellular, which involves transport as well as phosphoribosylation. Cytosoluble and membrane-bound APRTase and PRibPP synthetase exhibited different catalytic properties and sensitivities to effectors. Membranes of erythrocytes of HPRTase-deficient patients contain little or no HPRTase activity when assayed in the absence of Triton. Reisolation of these membranes from admixture with normal hemolysates did not result in any bound activity; thus, the membrane-bound activity is not an artifact of the isolation procedure. Lysis with Triton released activity equal to about half that of control membranes. This is further evidence that the enzyme is firmly bound to the membrane.
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PMID:Membrane-associated purine metabolizing enzyme activities of human peripheral blood cells. 629 41

Specific recognition of duplex DNA by a single-stranded oligonucleotide via the formation of triplex DNA is a rational approach for targeting specific regions of a genome. By screening a number of potential target sites for triple helix formation within mammalian genes that allow genetic selection in cell culture, we have identified a site within intron 1 of the hamster adenine phosphoribosyltransferase (APRT) gene that specifically binds a triplex-forming oligodeoxyribonucleotide (TFO) with high affinity. Under optimal conditions for triplex formation, the equilibrium dissociation constant is in the nanomolar range (Kd = 7 x 10(-10) M). This high-affinity binding is very specific, as a 10(5)-fold excess of genomic DNA reduced triplex formation less than 10-fold, and within a 6928-bp plasmid bearing the APRT gene, only restriction fragments containing the intron 1 site were found to bind the TFO. Results of DNase I protection assays were consistent with the TFO binding in an antiparallel orientation via reverse Hoogsteen hydrogen bonds in the major groove of the duplex. We have examined the kinetics of triplex formation as well as the effects of ionic composition and chemical modifications of the TFO on triplex formation. While divalent cations were not required for triplex formation, Mg2+ stabilized the triplex apparently through inhibition of TFO dissociation, with a mean bound lifetime of > 17 h for the triplex at Mg2+ concentrations above 5 mM.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:High-affinity triple helix formation by synthetic oligonucleotides at a site within a selectable mammalian gene. 776 35

The adenine phosphoribosyltransferase (APRTase) from Giardia lamblia was co-crystallized with 9-deazaadenine and sulfate or with 9-deazaadenine and Mg-phosphoribosylpyrophosphate. The complexes were solved and refined to 1.85 and 1.95 A resolution. Giardia APRTase is a symmetric homodimer with the monomers built around Rossman fold cores, an element common to all known purine phosphoribosyltransferases. The catalytic sites are capped with a small hood domain that is unique to the APRTases. These structures reveal several features relevant to the catalytic function of APRTase: 1) a non-proline cis peptide bond (Glu(61)-Ser(62)) is required to form the pyrophosphate binding site in the APRTase.9dA.MgPRPP complex but is a trans peptide bond in the absence of pyrophosphate group, as observed in the APRTase.9dA.SO4 complex; 2) a catalytic site loop is closed and fully ordered in both complexes, with Glu(100) from the catalytic loop acting as the acid/base for protonation/deprotonation of N-7 of the adenine ring; 3) the pyrophosphoryl charge is neutralized by a single Mg2+ ion and Arg(63), in contrast to the hypoxanthine-guanine phosphoribosyltransferases, which use two Mg2+ ions; and 4) the nearest structural neighbors to APRTases are the orotate phosphoribosyltransferases, suggesting different paths of evolution for adenine relative to other purine PRTases. An overlap comparison of AMP and 9-deazaadenine plus Mg-PRPP at the catalytic sites of APRTases indicated that reaction coordinate motion involves a 2.1-A excursion of the ribosyl anomeric carbon, whereas the adenine ring and the 5-phosphoryl group remained fixed. G. lamblia APRTase therefore provides another example of nucleophilic displacement by electrophile migration.
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PMID:Closed site complexes of adenine phosphoribosyltransferase from Giardia lamblia reveal a mechanism of ribosyl migration. 1217 25