Replication fork movement
During initiation to elongation transition, DnaB (while travelling in 5'-3' direction) generates a replication fork by opening the DNA duplex (Fig. 26.21). The strand having DnaB will thus be the lagging strand template. DNA primase associates with DnaB and synthesizes multiple primers for lagging strand and single primer for leading strand. The dimeric DNA pol III HE interacts simultaneously with both template strands. The leading strand polymerase moves in the same direction as DnaB, while copying the opposite strand. For the synthesis of lagging strand, the DNA polymerase will have to work on the same strand to which DnaB is bound, but it travels in opposite direction. The helicase and primase activities may be provided by DnaB and primase or alternatively by ΦX-type primosome.
|Fig. 26.21. Model of a DNA replication fork in prokaryotes during elongation.
DnaB, primase and 'DNA pol III HE' function together in elongation, although DnaB and DNA pol III HE seem to be tightly bound to the fork and stay bound throughout the reaction, an attribute described by saying that DnaB and DNA polymerase assembly is processive. In contrast, primase, 'β' and '
λcomplex' act distributively (they dissociate and reassociate i.e. do not stay bound throughout the reaction).
Discontinuous synthesis on lagging strand.
The events on lagging strand are more complex than on leading strand, and DNA polymerase enzyme is less processive. Primase is taken up from solution and activated by DnaB, to synthesize a primer (10-20 nt in length) on the lagging strand. The 'DNA pol III
HE' interacts with primase and provides the signal for primase to dissociate from the fork. The primers are recognized by the polymerase on the lagging strand and are utilized for synthesis of Okazaki fragments. The new primer is actually recognized by a 'y complex' and loaded with 'β subunit' (processivity factor). This 'preloaded β subunit' may then capture the polymerase (DNA pol III
core), when it becomes available after finishing synthesis of the preceding fragment.
Continuous synthesis on leading strand.
The leading strand is primed only once on
each of the two parental strands in bidirectional replication. In bidirectional replication, the first primer synthesized can serve as the 'leading strand primer' and subsequent primers serve as 'lagging strand primers'. No special mechanism is, therefore, needed for priming of the leading strand in bidirectional replication, although the DNA polymerase complex on leading strand is extremely processive. However, in unidirectional replication an alternative mechanism for priming is needed for leading strand. For ColE1-type plasmids, RNA polymerase performs this role.