ACTIVE CENTER OF DNA POLYMERASE

DNA polymerase, a homogeneous protein of molecular weight 109,000, appears to be a single polypeptide chain. The enzyme contains one triphosphate substrate binding site and one site for binding a nicked region of duplex DNA. A model of the active center of the enzyme has been proposed (Fig. 11) in which there are distinctive sites for the template strand, the primer strand, the 3′-hydroxyl primer strand terminus, the triphosphate substate, and the 5′-phosphateterminated strand beyond the nick (point of scission). The model attempts to account for the various synthetic and degradative functions within the closely related sites in the active center of the enzyme. Initiation of replication is favored at a nicked region of a duplex. In the first phase of replication, extension of the 3′-hydroxyl primer strand appears to be related to the 5′ → 3′ hydrolytic removal of the 5′-phosphate-terminated strand. The failure of phage-induced DNA polymerases to initiate replication at a nicked region may be due to the lack of a 5′ → 3′ nuclease function in the phage enzyme. A speculative model for helix replication, in vivo (Fig. 13), suggests how DNA polymerase, in conjunction with endonuclease and ligase, may achieve the sequential and almost simultaneous replication of both strands of a helix.