Synthesis and biological activity of N6-(N[(4-azido-3,5,6-trifluoro)-pyridin-2-yl]-2-aminoethyl)-adenosine 5'-monophosphate, a new AMP photoactivatable derivative. Covalent modification of horse liver alcohol dehydrogenase.
N6-(N-[(4-Azido-3,5,6-trifluoro)pyridin-2-yl]-2-aminoethyl)- adenosine 5'-monophosphate has been synthesized and evidence presented for its structural assignment by ultraviolet and 19F-NMR spectroscopies. Its photolysis was shown to occur within 5 min. This AMP derivative behaves as a competitive inhibitor of NAD+ in horse-liver-alcohol-dehydrogenase-promoted oxidation of ethanol, with a Ki (0.95 mM) comparable to the Ki of AMP (1.9 mM). Moreover it is an activator of the enzyme when nicotinamide ribose is used as the oxidation cofactor. This activation is as good as that promoted by AMP or by the well known 8-azido-AMP. Upon photolysis of this new derivative in the presence of horse liver alcohol dehydrogenase, a covalent enzyme--analogue complex was isolated and assayed as a catalyst in the oxidation of ethanol using nicotinamide ribose as the cofactor. The reaction took place without complementation of AMP, indicating clearly that the AMP analogue is mainly covalently bound in the AMP-binding site, and that the linkage formed between the enzyme and the azido derivative has not dramatically altered the active site of the enzyme. A similar experiment with 8-azido-AMP produced a completely inactive complex.
Sicsic
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Centre d'Etudes et de Recherches de Chimie Organique Appliquée, Centre National de la Recherche Scientifique, Thiais, France.
Leonil
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J
Le Goffic
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F
D000249:Adenosine Monophosphate / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000494:pharmacology / Q000528:radiation effects; D000426:Alcohol Dehydrogenase / Q000378:metabolism* / Q000528:radiation effects; D000818:Animals; D001386:Azides / Q000138:chemical synthesis* / Q000494:pharmacology / Q000528:radiation effects; D001665:Binding Sites; D001667:Binding, Competitive; D002384:Catalysis; D008099:Liver / Q000201:enzymology*; D009682:Magnetic Resonance Spectroscopy; D015394:Molecular Structure; D009243:NAD / Q000037:antagonists & inhibitors; D010084:Oxidation-Reduction / Q000187:drug effects; D010777:Photochemistry; D010782:Photolysis; D013056:Spectrophotometry, Ultraviolet
D001386:Azides; D009243:NAD; C058663:N(6)-(N((4-azido-3,5,6-trifluoro)pyridin-2-yl)-2-aminoethyl)adenosine 5'-monophosphate; D000249:Adenosine Monophosphate; D000426:Alcohol Dehydrogenase