What enzyme catalyzes the formation of phosphodiester bonds during DNA ligation, and what cofactor is typically required?

• A. DNA polymerase; ATP.
• B. RNA ligase; NAD+.
• C. DNA ligase; ATP.
• D. Topoisomerase; GTP.

Answer: (C)

Sealing a nick in the DNA backbone requires a ligase that can create a new phosphodiester bond, using energy input to drive the reaction. DNA ligase performs this task by activating the 5' phosphate with ATP, forming a ligase–AMP intermediate, which then transfers the AMP to the 5' phosphate. This activated phosphate is then attacked by the 3' hydroxyl end to form the new phosphodiester bond and release AMP, effectively joining the DNA strands. In most organisms, this ATP-dependent mechanism is the standard way ligation is powered, which is why ATP is the typical cofactor. Some bacterial ligases do use NAD+ instead of ATP, but ATP-dependent ligation is the common scenario in eukaryotes and many contexts. Others listed—DNA polymerase, while it makes DNA bonds during synthesis, does not seal nicks in the same ligation step; RNA ligase specializes in RNA at times; and topoisomerase modulates DNA topology, not the standard nick sealing step.