How does proofreading by DNA polymerase contribute to mutation prevention?

• A. 5'→3' exonuclease proofreading removes mispaired nucleotides immediately after incorporation, improving fidelity before the next nucleotide is added.
• B. 3'→5' exonuclease proofreading removes mispaired nucleotides immediately after incorporation, improving fidelity before the next nucleotide is added.
• C. DNA polymerase proofreading occurs only after DNA replication is complete.
• D. Mismatch repair removes nucleotides after replication.

Answer: (B)

Proofreading by DNA polymerase relies on a 3'→5' exonuclease activity to correct mistakes right after a nucleotide is added, before the next one is incorporated. If the last added base is mismatched, the enzyme backtracks slightly, the incorrect nucleotide is removed from the 3' end, and then the correct nucleotide is inserted opposite the template, allowing synthesis to continue. This immediate correction greatly lowers the chance that an error will be fixed later or propagated into the daughter strands.

The alternative that suggests a 5'→3' exonuclease activity describes a different function (such as primer removal or processing) and is not the proofreading mechanism. Mismatch repair is a separate system that acts after replication to fix remaining errors, but it is not the proofreading performed by DNA polymerase during synthesis.