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The deoxyribonucleic acid polymerases from the diatom Cylindrotheca fusiformis. Partial purification and characterization of four distinct activities.

ABSTRACT: Four extramitochondrial DNA polymerases from the marine photosynthetic diatom Cylindrotheca fusiformis were isolated and purified more than 1200-fold by chromatography on DNA-cellulose and DEAE-Sephadex. The enzymes were equally susceptible to inhibition by the thiol-blocking agents N-ethylmaleimide and p-chloromercuribenzoate, the zinc chelator o-phenathroline, and the nucleic acid interchelators ethidium bromide and acriflavin; they displayed similar pH optima, preferred activated DNA, and had strict dependence on high K+ for maximum activity. They were differentiated on the basis of their kinetic parameters, template-primer utilization and salt requirements. The four activities varied with growth stage of C. fusiformis. Activities of polymerases A and D doubled in exponential-phase cells as compared with those in stationary-phase cells, and the increase in polymerase B and chloroplast activity C was 20-40%. The relationship of the diatom polymerases to the complements in other organisms is discussed.

SUBMITTER: Okita TW

PROVIDER: S-EPMC1183706 | biostudies-other | 1977 Dec

REPOSITORIES: biostudies-other

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Deoxyribonucleic acid polymerases of Euglena gracilis. Purification and properties of two distinct deoxyribonucleic acid polymerases of high molecular weight.

Project description: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.

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Four-Dimensional Deoxyribonucleic Acid-Gold Nanoparticle Assemblies.

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Deoxyribonucleic acid polymerases of Euglena gracilis. Primer-template utilization of and enzyme activities associated with the two deoxyribonucleic acid polymerases of high molecular weight.

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The deoxyribonucleic acid polymerases from the diatom Cylindrotheca fusiformis. Subcellular distribution, exonuclease activity and heterogeneity of the enzymes.

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