Characterisation of Nuclear and Cytoplasmic DNA Polymerases From Rat Intestinal Mucosa

Some properties of two DNA polymerases isolated from purified nuclei of rat intestinal mucosa were compared with those of a cytoplasmic DNA polymerase from the same tissue. The molecular weight of the NaCl-soluble nuclear polymerase was estimated to be 39 000, while a value of 100 000 was determined for both the Tris-soluble nuclear and cytoplasmic enzymes. The high molecular weight DNA polymerases shared many other common characteristics. Their chromatographic properties on DEAE-cellulose and phosphocellulose were identical. In the presence of 50 mM salt or 1 mM nalidixic acid the activities of the Tris-soluble nuclear and cytoplasmic enzymes were inhibited, whereas the activity of the low molecular weight NaCl-soluble nuclear enzyme was greatly enhanced. The activities of the three DNA polymerases were inhibited by sulfhydryl blocking agents. The NaCl-soluble nuclear enzyme showed maximal activity when native DNA that had been treated briefly with DNase I was supplied as the primer–template. Extensive treatment of the native DNA with DNase I, which converted 15% of it to acid-soluble form, reduced its templating activity by more than 90%. No incorporation was detected when denatured DNA was supplied as primer–template. In contrast, both native and denatured DNA served as primer–templates for the Tris-soluble nuclear and cytoplasmic DNA polymerases, and the most efficient primer–template tested was native DNA that had been rendered 15% acid soluble by digestion with DNase I.