1. The inhibitory effects of iodoacetate,
iodoacetamide,
p-hydroxymercuribenzoate and sarkomycin were studied in a partially purified preparation of
deoxyribonucleic acid nucleotidyltransferase from Landschutz
ascites-tumour cells. All of these agents inhibited the activity of the
enzyme, and it was shown that the inhibition exerted by the last three compounds obeyed non-competitive kinetics. 2. Inclusion of
glutathione or
2-mercaptoethanol in the
enzyme assays did not prevent the inhibition by iodoacetate or
iodoacetamide, but did prevent inhibition by
p-hydroxymercuribenzoate. Inhibition by sarkomycin could be partially prevented by
glutathione or
2-mercaptoethanol. 3. The
enzyme fraction also catalysed incorporation in the presence of only one
triphosphate (thymidine 5'-triphosphate), and the limited incorporation observed in these circumstances was more resistant to the inhibitory action of
iodoacetamide and p-hydroxy-mercuribenzoate than was the standard
nucleotidyltransferase reaction (four triphosphates present). Levels of inhibition imposed on the standard reaction were achieved in the limited incorporation reaction with 2.5-fold higher concentrations of the two inhibitors. 4. The addition of certain bivalent
cations to the standard system resulted in severe inhibition of the reaction: Zn(2+)
ions (10mum) gave 50% inhibition; ethylenediaminetetra-
acetate (0.4mm) in the reaction mixture gave essentially complete protection against this inhibitory effect of Zn(2+)
ions. 5.
Deoxyribonucleic acid-
nucleotidyltransferase fractions prepared in the presence of a
thiol and ethylenediaminetetra-
acetate could be stored without loss of activity for 2 months at 0 degrees or for 1 year at -70 degrees .