DNA polymerase-beta from the nuclear fraction of sea urchin embryos: Characterization of the purified enzyme

• Published in: Journal of biochemistry (Tokyo) Vol. 82; no. 6; pp. 1613 - 1621
• Main Authors: Suzuki-Hori, C, Nagano, H, Mano, Y
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 01.01.1977
• Subjects: Animals; Cations, Divalent; Cell Nucleus - enzymology; DNA Polymerase II - isolation & purification; DNA Polymerase II - metabolism; DNA-Directed DNA Polymerase - metabolism; Embryo, Nonmammalian; Kinetics; Sea Urchins - enzymology; Sulfhydryl Reagents - pharmacology
• ISSN: 0021-924X; 1756-2651
• DOI: 10.1093/oxfordjournals.jbchem.a131857

Approximately 2,500-fold purification of DNA po1ymerase-β from the nuclear fraction of blastulae of the sea urchin, Hemicentrotus pulcherrimus, was performed. The enzyme prepa ration, which was devoid of DNase and terminal deoxynucleotidyl transferase as contaminants, showed a sedimentation constant of 3.0 S in a sucrose density gradient, a molecular weight of 50,000 by gel filtration, and an isoelectric point of pH 8.1. The enzyme activity was resistant to sulfhydryl group inhibitors. Its optimal pH was 9.0–9.5 in Tris-maleate buffer and 10.0 in glycine buffer. The optimal NaCl concentration for the activity was 30–60 nmt and about half of the activity remained at 0.4 M NaCl. As a template-primer, the enzyme preferred synthetic homopolymers to activated DNA. The order of this preference was as follows; poly (dA)-oligo (dT)12–16 > poly (rA)-oligo (dT)12–16 > activated DNA. The above results indicate that the enzyme corresponds to DNA polymerase-β from vertebrate cells.