Purification and properties of two growth stage-dependent deoxyribonucleic acid polymerases from Lactobacillus acidophilus.

Two distinct deoxyribonucleic acid polymerase activities have been purified from extracts of Zactobacillus acidophilus. Fraction I may be prepared from log phase and stationary phase cells. Fraction II appears initially in late log phase and increases to a maximum of 4.5 % of the total DNA polymerase activity found in extracts of stationary phase cells. Fraction I and Fraction II have been purified approximately 1000-fold and 35O-fold, respectively. Sedimentation coefficients of the two enzymes were 5.2 f 0.1 S (78,000 molecular weight) for Fraction I and 5.8 & 0.1 S (92,000 molecular weight) for Fraction II. Although the pH optima and magnesium titration curves for the two fractions were similar, several significant differences in their catalytic behavior were observed. Fraction I used either native or denatured primertemplate DNA from Z. acidophilus with equal efficiency. The addition of spermidine caused a 4-fold stimulation in activity of Fraction I assayed on denatured DNA. Fraction II showed a preference for native DNA in the absence of spermidine and used native or denatured DNA equally well with spermidine present. Fraction I was much less sensitive to inactivation by parahydroxymercuribenzoate than Fraction II, and the two fractions had dissimilar manganese titration curves. Thermal inactivation curves at 45” provided further evidence that the two fractions were different, and similar studies with mixed extracts suggested that the two polymerases may interact with each other.