Studies on the purification and properties of a 6.8-S DNA polymerase activity found in calf-thymus DNA polymerase-alpha fraction

Eur J Biochem. 1978 Mar;84(1):123-31. doi: 10.1111/j.1432-1033.1978.tb12148.x.

Abstract

The heterogeneity of calf thymus DNA polymerase-alpha has been further investigated. In particular, an enzyme (enzyme D) which exhibits higher activity on poly(dA) . (dT)10 (A:T = 20:1) compared with that on activated DNA, has been further purified and its properties compared with two other activities of the DNA polymerase-alpha fraction (enzymes A1 and C) which do not show a preference for poly(dA) . (dT)10 over activated DNA. As with A1 and C, enzyme D was shown to have many of the characteristic properties of DNA polymerase-alpha in that it is an acidic protein as judged by its binding to DEAE-cellulose, has a molecular weight of about 140000, does not use a poly (A) . (dT)10 template-initiator complex and is inhibited by N-ethylmaleimide. It exhibits anomalous gel filtration behaviour on Sepharose 6B and it binds relatively weakly to DNA-cellulose compared with DNA polymerase-beta. The extreme sensitivity of enzyme D to inhibtion by N-ethylmaleimide distinguishes it from A1 and C, as does its elution position from a DEAE-cellulose column. On the other hand enzymes C and D are readily inactivated by heating at 45 degrees C unlike enzyme A1. The possible interrelationships of the multiple activities of calf thymus DNA polymerase-alpha are discussed.

MeSH terms

  • Animals
  • Cattle
  • DNA Polymerase II / metabolism
  • DNA-Directed DNA Polymerase / isolation & purification
  • DNA-Directed DNA Polymerase / metabolism*
  • Ethylmaleimide / pharmacology
  • Hot Temperature
  • Hydrogen-Ion Concentration
  • Liver / enzymology
  • Molecular Weight
  • Poly dA-dT / metabolism
  • Rats
  • Templates, Genetic
  • Thymus Gland / enzymology*

Substances

  • Poly dA-dT
  • DNA Polymerase II
  • DNA-Directed DNA Polymerase
  • Ethylmaleimide