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Control of initiation at oriC by methylation

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  • A new round of replication will initiate on the E. coli chromosome at oriC only when the growth conditions permit it. The dam methylase and features of its sites of action are used to prevent premature re-initiation. The dam methylase of E. coli recognizes the tetranucleotide GATC in DNA and transfers a methyl group (from S‑adenosyl methionine) to the amino group at position 6 of the adenine in that sequence. Note that GATC is a pseudopalindrome, so both strands read the same for these four nucletides in DNA.

    Palindrome and Pseudopalindromes

    A nucleotide sequence is said to be a palindrome if it has an even number of base pairs and is equal to the reverse of its complementary sequence. For example, in a single strand of DNA the sequence of bases CCATTAATGG is palindromic because the sequence of bases in the complementary strand is GGTAATTACC, its reverse.

    A pseudopalindrome is a DNA sequence with an odd number of base pairs yielding a symmetrical complement except at the central base-pair. For example, the DNA sequence ACCTGGT is pseudopalindromic, because its complement on the other strand is TGGACCA, which is its reverse except for the central element.

    Thus a GATC in duplex DNA can be unmethylated on either strand, methylated on only one strand (referred to as hemimethylated) or methylated on both strands (referred to as fully methylated), as shown in Figure \(\PageIndex{1}\).

    Figure \(\PageIndex{1}\)

    The methylation status of the 11 GATC motifs at oriCregulate whether replication can intitiate. When the GATCs are fully methylated, oriCDNA serves as an origin (in the presence of Dna A and the other proteins discussed above). However, when the GATCs are hemimethylated, it is not active as an origin. The reason for this is not fully known. One hint comes from the behavior of unmethylated oriC(from dam- strains). This unmethylated oriC is active, showing that methylation of the GATC is not a requirement for initiation, and further suggesting that some inhibitor of initiation recognizes the hemimethylated form.

    Exercise \(\PageIndex{1}\)

    How do these results lead to this conclusion? Let’s explore this by posing the opposite hypotheses.

    1. If methylation of the GATC motifs at oriC were needed for initiation, what would the result have been?
    2. If some activator recognized the fully methylated form, what would the result have been.?

    Re-methylation of oriCby the dam methylase is quite slow. Thus for some period the GATCs at oriCare hemimethylated, and the origin is inactive. This provides a means to delay the use of oriCto start another round of replication. Thus the methylation of the GATCs is part of a mechanism to regulate the timing of firing of oriC. In the next chapter, we will also see the use of methylation of GATCs in post-replicative repair.