INACTIVATION OF DNA-POLYMERASE ALPHA-PRIMASE BY ACROLEIN - LOSS OF ACTIVITY DEPENDS ON THE DNA SUBSTRATE

We have utilized acrolein as a model compound to examine the biochemic al behavior of chemically-modified DNA polymerase alpha-primase comple x (pol alpha). We have found that acrolein irreversibly inactivates th e DNA synthetic capacity of pol alpha polymerase in a time- and concen tration-dependent manner. Double stranded DNA protects pol alpha polym erase from inactivation when present during acrolein exposure, but sin gle-stranded DNA, dATP and ATP do not. Strikingly, the activity of pol alpha polymerase is strongly dependent upon the DNA substrate utilize d to assay catalytic activity after exposure to the aldehyde. The prim ase activity of pol alpha is also inactivated by exposure to acrolein, but the observed rate of inactivation is slower than that seen for DN A synthesis. Competitive labeling studies with [C-14] iodoacetamide su ggest that acrolein inactivation of the enzyme is mediated through the modification of protein sulfhydryl groups. (C) 1995 Academic Press, I nc.