passages: PMC-1635332-sec-23__text
passages: sec
passages: Nf and GA-1 DNA polymerases couple polymerization to strand displacement processively
  The results presented in this paper clearly indicate that Nf and GA-1 DNA polymerases can account for their genome replication without the assistance of unwinding and processivity factors, in contrast to most replicative DNA polymerases which require their physical association to processivity factors and DNA unwinding proteins (1,70). Strand displacement capacity has also been shown for other protein-primed DNA polymerases as those of bacteriophages phi29 (10), Cp-1 (71) and PRD1 (72,73). On the contrary, adenovirus DNA polymerase, although processive, cannot couple polymerization to strand displacement, requiring the DNA unwinding activity of the adenovirus DBP to perform strand displacement (74,75).
  Whereas it was possible to obtain GA-1 DNA replication by using exclusively the GA-1 TP and DNA polymerase, Nf DNA polymerase, although provided with competent strand displacement and processivity features, required the presence of Nf DBP for an effective in vitro replication of Nf TP-DNA. Results presented here show that Nf DBP strongly stimulates the formation of the TP-dAMP initiation complex by decreasing the Km for dATP and facilitates the transition from initiation to elongation, as it occurs in phi29 (76). These results point to either a specific and direct contact between DBP and DNA polymerase that promotes conformational changes at the polymerization active site or to an effect of DBP in conferring the optimal template structure to direct initiating nucleotide insertion. A similar role has been proposed for adenovirus DBP, a DNA unwinding protein (77). As in the case of phi29 and Nf DBP, this protein stimulates the rate of initiation also by decreasing the Km for the initiating nucleotide (74). The fact that an adenovirus DBP mutant defective in unwinding can still stimulate initiation precludes the unwinding role as the one responsible for such an activation (77,78). In this case, contacts between DBP and pTP/DNA polymerase complex have been reported (77).
  The effect of Nf DBP in promoting elongation of the initiation products could be due to a decrease of the Km also for the incorporation of the dNMPs during the transition stage from initiation to elongation, to a different type of contact with the DNA polymerase that helps transition to elongation, or both. The similarity in replication rates when comparing M13 DNA replication, performed in the absence of DBP (2400 nt/min), with Nf TP-DNA replication in the presence of DBP (2260 nt/min), suggests that the DBP stimulatory role is restricted to the first phases of Nf TP-DNA replication.
  