Two Distinct Poly(A) Polymerases Isolated from the Cytoplasm of Ehrlich Ascites Tumour Cells

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Abstract
The poly(A) polymerases from the cytosol and ribosomal fractions of [mouse] Ehrlich ascites tumor cells were isolated and partially purified by DEAE-cellulose and phosphocellulose column chromatography. Two distinct enzymes were identified: a cytosol Mn2+-dependent poly(A) polymerase (ATP:RNA adenyltransferase) and a ribosome-associated enzyme defined tentatively as ATP(UTP):RNA nucleotidyltransferase. The cytosol poly(A) polymerase was strictly Mn2+-dependent (optimum at 1 mM Mn2+) and uses only ATP as substrate; poly(A) was a better primer than rRNA. The purified enzyme was free of poly(A) hydrolase activity, but degradation of [3H]poly(A) took place in the presence of PPi. Most likely this enzyme is of nuclear origin. The ribosomal enzyme was associated with the ribosomes, but it was also found in free state in the cytosol. The purified enzyme used both ATP and UTP as substrates. The substrate specificity varied depending on ionic conditions: the optimal enzyme activity with ATP as substrate was at 1 mM Mn2+, while that with UTP as substrate was at 10-20 mM Mg2+. The enzyme used both ribosomal RNA and poly(A), but not poly(U), as primers. The purified enzyme was free of poly(A) hydrolase activity.