Enzymatic preparation of 32P-labeled β-L-2′,3′-dd-5′ATP and its use as a high-affinity, conformation-specific ligand for labeling adenylyl cyclases

An enzymatic method was developed for the preparation of unlabeled and [beta- P-32]-labeled beta-L-2',3'-dd-5'ATP from the monophosphate with near quantitative yields. beta-L-2',3'-dd-5'ATP was a competitive and potent inhibitor of adenylyl cyclases (IC5 similar to 30 nM). Upon uv-irradiation beta-L2',3'-dd-[beta-(32)p]-5'ATP directly crosslinked to a chimeric construct of this enzyme. Data suggest that this is a pre-transition state inhibitor and contrasts with the equipotent 2',5'-dd-3'ATP, a post-transition state, noncompetitive inhibitor.