10 S DNA polymerase alpha of calf thymus shows a microheterogeneity in its large polypeptide component

J Biol Chem. 1982 Jun 25;257(12):7172-7.

Abstract

The 10 S DNA polymerase alpha from calf thymus (Masaki, S., and Yoshida, S. (1978) Biochim. Biophys. Acta 521, 74-88) has been purified to near homogeneity. The most purified fraction obtained by repeated sucrose rate-zonal centrifugation contained three large polypeptides of 150,000, 145,000, and 140,000 daltons and three to four smaller polypeptides ranging from 43,000 to 50,000 daltons. A good resolution of these polypeptides was achieved on a sodium dodecyl sulfate-polyacrylamide linear gradient gel (5-20%) which was stained by the silver stain method. The three large polypeptides were also observed in the more crude fractions prepared in the presence of three kinds of protease inhibitors. By a peptide mapping analysis, it was revealed that these three polypeptides have a similar primary structure. Treatments of the enzyme with alkaline phosphatase, phosphodiesterase, and neuraminidase did not affect the gel pattern. These results indicate that the 10 S DNA polymerase alpha of calf thymus has a microheterogeneity in terms of the large polypeptide component. Among these three large polypeptides, the two polypeptides of 150,000 and 145,000 daltons disappeared by keeping the sucrose gradient fraction at 4 degrees C in the absence of glycerol, while the 140,000-dalton polypeptide was well preserved. The poly(rA)oligo(dT)-dependent activity of 10 S DNA polymerase alpha was selectively lost under this condition.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cattle
  • DNA Polymerase II / isolation & purification*
  • DNA Polymerase II / metabolism
  • DNA-Directed DNA Polymerase / isolation & purification*
  • Drug Stability
  • Glycerol / pharmacology
  • Kinetics
  • Macromolecular Substances
  • Molecular Weight
  • Peptide Fragments / analysis
  • Thymus Gland / enzymology*

Substances

  • Macromolecular Substances
  • Peptide Fragments
  • DNA Polymerase II
  • DNA-Directed DNA Polymerase
  • Glycerol