RESUMO
A one step synthesis of fluorescent 8-aryl-(7-deazaguanines) has been accomplished. Probes exhibit blue to green high quantum yield fluorescence in a variety of organic and aqueous solutions, high extinction coefficients, and large Stokes shifts often above 100 nm. The probes are highly cell permeable, and exhibit stable bright fluorescence once intracellular; therefore are suited to the design of biosensors.
Assuntos
Azaguanina/química , Azaguanina/metabolismo , Permeabilidade da Membrana Celular , Corantes Fluorescentes/química , Corantes Fluorescentes/metabolismo , Azaguanina/análogos & derivados , Azaguanina/síntese química , Linhagem Celular Tumoral , Fluorescência , Corantes Fluorescentes/síntese química , Humanos , Células KB , Microscopia Confocal , Estrutura MolecularRESUMO
Various forms of purine-nucleoside phosphorylase (PNP) were used as catalysts of enzymatic ribosylation of selected fluorescent 8-azapurines. It was found that the recombinant calf PNP catalyzes ribosylation of 2,6-diamino-8-azapurine in a phosphate-free medium, with ribose-1-phosphate as ribose donor, but the ribosylation site is predominantly N7 and N8, with the proportion of N8/N7 ribosylated products markedly dependent on the reaction conditions. Both products are fluorescent. Application of the E. coli PNP gave a mixture of N8 and N9-substituted ribosides. Fluorescence of the ribosylated 2,6-diamino-8-azapurine has been briefly characterized. The highest quantum yield, ~0.9, was obtained for N9-ß-d-riboside (λmax 365 nm), while for N8-ß-d-riboside, emitting at ~430 nm, the fluorescence quantum yield was found to be close to 0.4. Ribosylation of 8-azaguanine with calf PNP as a catalyst goes exclusively to N9. By contrast, the E. coli PNP ribosylates 8-azaGua predominantly at N9, with minor, but highly fluorescent products ribosylated at N8/N7.
Assuntos
Azaguanina/análogos & derivados , Azaguanina/síntese química , Proteínas de Escherichia coli/química , Corantes Fluorescentes/síntese química , Purina-Núcleosídeo Fosforilase/química , Animais , Biocatálise , Bovinos , Glicosilação , Cinética , Proteínas Recombinantes/química , Ribosemonofosfatos/químicaRESUMO
Phosphonate acyclic derivates of guanines, pyrazolo[3,4-d]pyrimidines, and triazolo[4,5-d]-pyrimidines (8-azaguanines) are inhibitors of the enzyme purine nucleoside phosphorylase (PNPase) with Ki' values ranging from 0.05 to 1.6 microM. These compounds are enzymatically stable congeners of the potent PNPase inhibitor acyclovir diphosphate (53).