title: Synthesis of a new 8-spin-labeled analog of adenosine 5'-phosphate and its interaction with AMP nucleosidase.
abstract: The synthesis of a new 8-spin-labeled analog of AMP, 8-[[[(2,2,5,5-tetramethyl-1-oxy-3-pyrrolidinyl)carbamoyl]methyl]thio]adenosine 5'-phosphate (8-slAMP), is described. The procedure is facile and results in high yields. 8-slAMP is a competitive inhibitor of AMP nucleosidase with a Ki of 19 microM as compared to a Km of 100 microM for AMP. The analog is not a substrate for the enzyme and does not displace MgATP2- from the allosteric sites under the usual assay conditions. The EPR spectrum of the bound spin probe reveals a highly immobilized nitroxide group. Binding studies with 8-slAMP at 8 degrees C indicate three independent binding sites (Kd = 1.4 microM) per molecule of enzyme (Mr = 320,000). These properties make 8-slAMP a good spin probe for AMP nucleosidase. The analog may also be useful for other proteins known or suspected of binding AMP analogs in a syn conformation.
mesh_terms: D000249:Adenosine Monophosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D001395:Azotobacter / Q000201:enzymology; D001665:Binding Sites; D004578:Electron Spin Resonance Spectroscopy; D007700:Kinetics; D009699:N-Glycosyl Hydrolases / Q000378:metabolism; D011485:Protein Binding; D013113:Spin Labels
chemical_list: D013113:Spin Labels; D000249:Adenosine Monophosphate; D009699:N-Glycosyl Hydrolases; C019699:AMP nucleosidase