Synthesis and use of 8-azidoguanosine 35-cyclic monophosphate as photoaffinity label for cyclic GMP-dependent protein kinase.

摘要

8-Azidoguanosine 3',5'-cyclic monophosphate (8-N3cGMP) has been synthesized for use as a photoactive probe for the labeling of cGMP receptors. The ability of 8-N3cGMP to be bound at specific cGMP binding sites was demonstrated by its ability to activate cGMP-dependent protein kinase isolated from bovine lung (Ka = 1.1 x 10(-7) M) and to inhibit competitively the binding of [3H] cGMP to the enzyme [Kd (8 N3GMP)/Kd (cGMP) = 6]. Photolysis of 8-N3[32P]cGMP in the presence of a crude enzyme preparation resulted in the covalent attachment of analog to cGMP-dependent protein kinase. Half-maximal labeling occurred at 2.2 x 10(-7) M. The incorporation of the analog was completely inhibited by the addition of cGMP.