Replication protein A (RPA), a three-subunit SSB complex binds to newly created single-stranded regions, to complete the formation of a proto-replication bubble. RPA prevents single strands from either reassociating or forming intrastrand secondary structures and also interacts with T antigen. RPA and T antigen help to recruit DNA polymerase α-primase, a four-subunit protein complex, to the proto-replication bubble. As the only eukaryotic enzyme that can synthesize primer Pol α is an essential participant in initiation. It is crystal structure has not yet been determined. Pol αhas two distinct catalytic activities, primase and DNA polymerase, which reside in separate subunits.
The other two subunits appear to play important structural roles but no enzyme activities have as yet been assigned to them. Mammalian Pol α does not have proofreading activity. Primase activity synthesizes RNA oligomers that are 8 to 10 nucleotides long, which serve as primers for DNA synthesis. Then the weakly processive DNA polymerase that is associated with Pol α incorporates 15 to 25 deoxynucleotides before dissociating from the DNA template. Thus, the oligonucleotide synthesized by Pol α consists of an RNA primer joined to a short DNA segment. The first pair of initiator DNA molecules, which are formed on either side of the core origin, serve to prime leading strand formation. Initiator DNA molecules formed by Pol αlater in the replication process prime lagging strand synthesis.