Replication in bacteria and in eukaryotes is licensed and permitted to occur only once per cell cycle. Each replicon is allowed to fire only once.

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#Unit 3. Fundamental Processes

AT base pairs called the DNA unwinding element (DUE).

IHF (integration host factor) and FIS (factor for inversion stimulation). These two proteins were discovered as required components of certain recombination reactions.

Another DNA-binding protein, HU (a histone like bacterial protein originally dubbed factor U), also participates but does not have a specific binding site.

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#Unit 3. Fundamental Processes

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#Unit 3. Fundamental Processes

The single replicator 245 bp required for E. coli chromosomal replication is called oriC. Two repeated motifs are critical for oriC function.

The 9-mer motif is the binding site for the E. coli initiator, DnaA, and is repeated five times at oriC.

The 13-mer motif, repeated three times, is the initial site of ssDNA formation during initiation.

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#Unit 3. Fundamental Processes

oriC also contains 11 GATC/CTAG repeats that are methylated on adenine on both strands.

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#Unit 3. Fundamental Processes

Before replication, the palindromic target GATC site is methylated by dam methylase on the N6 of A of each strand.

Replication generates hemimethylated DNA, in which one strand is methylated and one strand is unmethylated.

Thus, the replication event converts Dam target sites from fully methylated to hemimethylated condition.

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#Unit 3. Fundamental Processes

Dam (Dam-) mutants accumulates hemimethylated products after replication.

If the plasmid is methylated, it undergoes a single round of replication, and then the hemimethylated products accumulate.

Suggesting that a hemimethylated origin cannot be used to initiate a replication cycle.