Heterogeneous forms of poly(rA) . oligo(dT)-directed DNA polymerase activity from rat spleen.
Three forms of DNA polymerase, named enzymes A, B, and C, that preferred (rA)n x (dT)12-18 as a template-primer, were partially purified from an extract of rat spleen. Enzymes B and C, both sedimenting at 9S, appeared to correspond to DNA polymerase gamma. However, they differed in their behavior on phosphocellulose and DNA-cellulose column chromatographies, and in their optimum KCl and divalent cation requirements for activity. Enzyme A showed a unique property. Like DNA polymerase beta, it sedimented at 3.8S, was resistant to reagents blocking sulfhydryl groups, and was inhibited by phosphate, but it differed from DNA polymerase beta with respect to elution positions from DEAE-cellulose, phosphocellulose and DNA-cellulose columns, Km value (lower by one order of magnitude for dTTP), and template-primer preference. Enzyme A was found in the mitochondrial fraction, in which DNA polymerase beta was not detectable. Enzymes A and C were isolated from the nuclear fraction, but this fraction did not contain enzyme B. The cytosol contained only enzyme A. The mitochondrial fraction contained enzyme A and enzyme C-like polymerase. Enzyme B was obtained with enzymes A and C only by extraction of the whole cell homogenate. Enzyme B may be labile or may be an artificial form of DNA polymerase gamma formed during the purification procedures.
D000818:Animals; D004259:DNA-Directed DNA Polymerase / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008970:Molecular Weight; D010710:Phosphates / Q000494:pharmacology; D011061:Poly A; D011071:Poly T; D011189:Potassium Chloride / Q000494:pharmacology; D051381:Rats; D013154:Spleen / Q000201:enzymology*; D013347:Subcellular Fractions / Q000201:enzymology; D013379:Substrate Specificity; D013439:Sulfhydryl Reagents / Q000494:pharmacology; D013698:Templates, Genetic
D010710:Phosphates; D013439:Sulfhydryl Reagents; D011061:Poly A; D011071:Poly T; D011189:Potassium Chloride; D004259:DNA-Directed DNA Polymerase