The image demonstrates how ligase (yellow oval) catalyzes two DNA fragment strands. The ligase joins the two fragments of DNA to form a longer strand of DNA by "pasting" them together.
The mechanism of DNA ligase is to form two covalent phosphodiester bonds between 3' hydroxyl ends of one nucleotide ("acceptor"), with the 5' phosphate end of another ("donor"). Two ATP molecules are consumed for each phosphodiester bond formed. AMP is required for the ligase reaction, which proceeds in four steps:
- Reorganization of activity site such as nicks in DNA segments or Okazaki fragments etc.
- Adenylation (addition of AMP) of a lysine residue in the active center of the enzyme, pyrophosphate is released;
- Transfer of the AMP to the 5' phosphate of the so-called donor, formation of a pyrophosphate bond;
- Formation of a phosphodiester bond between the 5' phosphate of the donor and the 3' hydroxyl of the acceptor.
A pictorial example of how a ligase works (with sticky ends
Ligase will also work with blunt ends, although higher enzyme concentrations and different reaction conditions are required.