Protection of Micrococcal Nuclease against Thermal Inactivation

Micrococcal nuclease in solution is protected from thermal inactivation for at least 5 hours by the simultaneous presence of 0.01 mCa++, 0.05% bovine serum albumin, and one of the products of the enzymatic reaction. The following products of digestion of DNA render protection: each of the four 3'-mononucleotides, those of deoxyadenosine and deoxythymidine being the most effective; and each of the tested dinucleotides, among which that of deoxyadenosine is the best and that of deoxyguanosine, the worst. When any of the three components (albumin, Ca++, or products) is omitted the other two are ineffective, suggesting that the mechanism of protection involves an interaction between all three and the enzyme. Each of the four 3',5'-deoxynucleoside diphosphates inhibits the action of micrococcal nuclease to some extent. However, the 3',5'-deoxynucleoside diphosphates of thymine and adenine are much more effective inhibitors than those of guanine and cytosine. The same type of base preference as that previously established for the hydrolytic action of the enzyme is now described for the protective effect of the reaction products and for the inhibiting effect of 3',5'-deoxynucleoside diphosphates. All three phenomena, therefore, appear to occur at, or in the vicinity of, the active center.