IDENTIFICATION AND PROPERTIES OF THE CATALYTIC DOMAIN OF MAMMALIAN DNA POLYMERASE-BETA

Rat DNA polymerase β (β-pol) is a 39-kDa protein organized in two tightly folded domains, 8-kDa N-terminal and 31-kDa C-terminal domains, connected by a short protease-sensitive region. The 8-kDa domain contributes template binding to the intact protein, and we now report that the 31-kDa C-terminal domain contributes catalytic activity. Our results show that this domain as a purified proteolytic fragment conducts DNA synthesis under appropriate conditions but the kcat is lower and primer extension properties are different from those of the intact enzyme. A proteolytic truncation of the 31-kDa catalytic domain fragment, to remove a 60-residue segment from the NH2-terminal end, results in nearly complete loss of activity, suggesting the importance of this segment. Overall, these results indicate that the domains of β-pol have distinct functional roles, template binding and nucleotidyltransferase, respectively; yet, the intact protein is more active for each function than the isolated individual domain fragment. © 1990, American Chemical Society. All rights reserved.