Deoxyribonucleic acid polymerases of Euglena gracilis. Purification and properties of two distinct deoxyribonucleic acid polymerases of high molecular weight.
Two DNA polymerases of high molecular weight, pol A (mol.wt. 190 000) and pol B (mol.wt. 240 ooo), have been purified 6300-fold and 1600-fold respectively from an extramitochondrial supernatant of a bleached strain of Euglena gracilis. They have very similar requirements when assayed with an 'activated'-DNA primer-template [the optimum conditions of pH and ionic (K+ and Mn2+) composition being 7.2, 25 mM and 0.2 mM respectively]. 0.2 mM-Mn2+ was about 1.5-2-fold as effective as 2 mM-Mg2+, owing to substrate activation by deoxyribonucleoside 5'-triphosphates in the presence of Mn2+. Km values for the triphosphates in the absence of activation were about 10(-6)M with Mn2+ and 8 X 10(-6) M with Mg2+ for both enzymes. They were inhibited to the same extent by N-ethylmaleimide, novobiocin and o-phenanthroline, but differed in their chromatographic behaviour on DEAE-cellulose and in their electrophoretic mobilities on polyacrylamide gel. No evidence was found for the existence in these cells of a DNA polymerase of low molecular weight, but there were indications that a third enzyme of high molecular weight might exist.
D002413:Cations, Divalent; D002414:Cations, Monovalent; D002845:Chromatography; D002847:Chromatography, Agarose; D002848:Chromatography, DEAE-Cellulose; D004254:DNA Nucleotidyltransferases / Q000032:analysis / Q000302:isolation & purification* / Q000378:metabolism; D003853:Deoxyribonucleosides / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005033:Ethylmaleimide / Q000494:pharmacology; D005056:Euglena gracilis / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000302:isolation & purification*; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D008970:Molecular Weight; D009675:Novobiocin / Q000494:pharmacology; D010618:Phenanthrolines / Q000494:pharmacology; D013347:Subcellular Fractions / Q000201:enzymology; D013438:Sulfhydryl Compounds / Q000494:pharmacology
D002413:Cations, Divalent; D002414:Cations, Monovalent; D003853:Deoxyribonucleosides; D007527:Isoenzymes; D010618:Phenanthrolines; D013438:Sulfhydryl Compounds; D009675:Novobiocin; D008345:Manganese; D004254:DNA Nucleotidyltransferases; D008274:Magnesium; D005033:Ethylmaleimide