Substrate specificity of a bacterial UV endonuclease and the overlap with in vitro photoanzymatic repair

Abstract— The action of an endonuclease from Micrococcus luteus , that operates on ultraviolet (UV) radiation damage, overlaps greatly with that of the yeast photoreactivating enzyme: homo and hetero cyclobutyl pyrimidine dimers in DNA are substrate for both enzymes, but pyrimidine adducts or the 'spore photoproduct' in DNA are not. As expected from this overlap, the action of the two enzymes is mutually interfering: single‐strand nicks introduced by the endonuclease effectively preclude photoreactivation; conversely, formation of a photoreactivating enzyme‐dimer complex can prevent nicking by the UV endonuclease. While complex formation between photoreactivating enzyme and dimers in UV‐endonuclease‐treated DNA is apparently normal, the light‐dependent repair step either fails to occur or proceeds at a very low rate. Hence, besides the requirement of a minimum chain length for the function of the photoreactivating enzyme, there is the additional restriction on the position of the dimer in a polynucleotide strand. Finally, rough approximations of the rate constants, k 1 and k 2 , for the UV endonuclease indicate that the in vitro UV‐endonuclease‐dimer complex is relatively unstable, with dissociation of the complex being more probable than hydrolysis of the phosphodiester bond.