HIGHLY SELECTIVE AFFINITY LABELING OF DNA-POLYMERASE ALPHA-PRIMASE FROM HUMAN PLACENTA BY REACTIVE ANALOGS OF ATP

Highly selective affinity labeling of a DNA-polymerase alpha-primase c omplex from human placenta by o-formylphenyl esters of ATP, ADP and AM P was performed in a two-step procedure in which a substrate analog at tached to the active center was elongated by radioactive ATP If the co valent attachment is performed in the presence of poly(dT) template, t he ATP esters modify selectively the delta subunit of the complex. If poly(dT) is added after the covalent binding of the reagent, both delt a and gamma subunits become labeled. With the o-formylphenyl ester of AMP the delta-subunit is modified. The ADP ester modifies both the del ta and gamma subunit in the presence and absence of template. It is sh own that formylphenyl ester of ATP is not the substrate in the reactio n of elongation catalyzed by primase. The data obtained suggest the bi nding site of initiating substrate to be located in the region of cont act of the two subunits of primase. The role of the template in the fo rmation of the active site is discussed.