Resolution of DNA polymerase-α-primase complex and primase free DNA polymerase α from embryonic chicken brain.

ABSTRACT

DNA polymerase-α from embryonic chicken brain was resolved on DEAEcellulose into 3 component activities that remained distinct upon rechromatography. Product formation by each activity required exogenously added template-primer DNA, all 4 deoxynucleoside triphosphates, and a divalent metal cation. Each form incorporated [3H]- dTTP or [3H]-dCTP into a high molecular weight product that was identified as DNA by its chromatographic behavior and its sensitivity to DNase. High ionic strength, Nethylmaleimide, and the polymerase-α-specific inhibitor aphidicolin inhibited each activity; the apparent Ki, value of aphidicolin was 3·0 μΜ in each case. Based on these results, the 3 activities were identified as multiple forms of DNA polymerase-α . Experiments using embryonic chicken brains of various ages indicated that polymerase-α1, and polymerase-α3 reached maximal activity in 9-day-old embryos, while polymerase-α2 activity was elevated at a slightly later developmental stage. Using poly (dC) as template, high primase activity was detected in polymerase-α1, fractions.