Question

Direct Repair Cells normally repair the lesion \(O^{6}\)-meG by directly transferring the methyl group to the protein \(O^{6}\) methylguanine-DNA methyltransferase. For the nucleotide sequence \(\mathrm{AAC}\left(O^{6}-\mathrm{meG}\right) \mathrm{TGCAC}\), with a damaged (methylated) G residue, what would be the sequence of both strands of double- stranded DNA resulting from replication in each of the situations listed? a. Replication occurs before repair. b. Replication occurs after repair. c. Two rounds of replication occur, followed by repair.

Short answer

a. Before repair: AACATGCAC. b. After repair: AACGTCGAC. c. Two rounds before repair: AACATGCAC repeating.

Step-by-step solution

Understanding the Scenario

In this exercise, we are considering a damaged DNA sequence where one of the guanine bases has been methylated at the O^6 position. This methylated guanine can pair with thymine instead of cytosine, leading to a mutation during replication if not repaired beforehand. This sequence needs to be analyzed to determine the resulting sequences after different replication scenarios without or with repair.

Replication Before Repair

Before repair, the methylated guanine pairs with thymine during replication. The given sequence is 5'-AAC(O^{6}-meG)TGCAC-3'. During replication, the methylated G will pair as if it were a T, so the complementary strand produced will be 3'-TTG(A)ACGTG-5', resulting in 5'-AACATGCAC-3' on one strand and 3'-TTGACGTG-5' on the other.

Replication After Repair

If replication occurs after repairing the methylated G back to a normal G, the sequence replicates correctly. The repair corrects the DNA sequence to 5'-AACGTCGAC-3'. The replication will then result in the complementary strand being correctly formed: 3'-TTGCAGTGT-5' and 5'-AACGTCGAC-3' on the other strand.

Two Rounds of Replication Followed by Repair

If two rounds of replication occur before the repair, the mutation caused by the methylated guanine will propagate. After the first replication, you have genetic information based on the methylated pairing (5'-AACATGCAC-3' and 3'-TTGACGTG-5'). After a second round of replication, these erroneous pairings will be perpetuated, producing: 5'-AACATGCAC-3' with 3'-TTGACGTG-5', and a new replication based on these creates 5'-AACATGCAC-3' and 3'-TTGACGTG-5'. Repair after this does not change the incorrect pairings already established in both new DNA molecules.

Key concepts

Direct Repair

Direct repair is a mechanism in which cells fix DNA damage directly at the site where it occurs. This method of repair is both efficient and highly specific. When a methyl group is added to position O^6 of a guanine base, it forms O^6-methylguanine (O^6-meG). This specific lesion can be recognized and repaired without the need to remove the damaged base and its surrounding nucleotides.
The repair enzyme, called O^6-methylguanine-DNA methyltransferase (MGMT), acts by transferring the methyl group from guanine to itself, effectively removing the damage. Since this molecule can only act once per molecule, it is not reusable, but it helps prevent mutations by restoring the guanine to its correct state. This form of repair is vital as O^6-meG can mispair with thymine, creating mutations if left unchecked.

Methylated Guanine

Methylated guanine is a form of damaged DNA caused by the addition of a methyl group at the O^6 position of guanine. This type of DNA lesion is a common result of exposure to various alkylating agents. These agents can be found in the environment, in the food we eat, and sometimes as a byproduct of cellular processes.
When guanine is methylated, it loses its ability to pair correctly with cytosine. Instead, O^6-methylguanine (O^6-meG) can mispair with thymine during DNA replication. This mispairing leads to a transition mutation from G:C to A:T in the DNA sequence, introducing a wrong base-pair segment in the newly replicated DNA strand. If these errors are not corrected by effective DNA repair mechanisms, they may result in permanent mutations in the genome.

DNA Replication

DNA replication is a fundamental process where the DNA molecule produces an identical copy of itself. This process is essential for cell division and involves unwinding the DNA double helix, followed by the synthesis of new complementary strands.
In normal scenarios, each base of the original DNA acts as a template for its complementary base pairing. Adenine pairs with thymine, while guanine pairs with cytosine. However, when a damaged base like O^6-methylguanine is present, the replication machinery might incorrectly pair bases, resulting in mutations. If O^6-meG goes unrepaired, it pairs with thymine instead of cytosine, creating a genetic error in the new DNA strand.
Thus, when replication happens before repair, these incorrect pairings become integrated into the DNA, making it crucial that DNA repair mechanisms function efficiently prior to replication.

Mutations in DNA

Mutations in DNA occur when there is an alteration in the sequence of nucleotides. Mutations can arise naturally through errors during DNA replication or through damage by external factors like UV radiation and chemicals.
One common type of mutation emerges from pairing errors, which might be introduced when DNA damage such as methylation is not repaired before replication. A substitution mutation, where a base is replaced by another, can happen if O^6-methylguanine incorrectly pairs with thymine instead of cytosine during replication. This specific error results in a G:C to A:T transition mutation.
Mutations can have various effects on organisms. Some might be silent, having no impact, whereas others can lead to diseases or inherited genetic disorders if they occur in vital genetic regions. Proper DNA repair systems are crucial as they minimize the impact of mutations, preserving genetic integrity.