Figure 1. Pathways leading to the formation of the substrate of DNA ligase. The horizontal lines represent DNA strands associated by hydrogen-bonded base pairs, which are represented by the short vertical lines. (a) The red asterisk and lines represent a damaged base that is removed to leave a gap in the duplex. A DNA polymerase fills (green line) the gap to leave a nicked DNA. (b). The red coloured portion of the upper strand is the RNA primer of an Okazaki fragment that is also removed and the gap filled to produce a nicked molecule. DNA ligase eradicates the discontinuity by synthesizing a phosphodiester bond. Reprinted from Subramanya et al. (1996). Cell 85: 607–615. Copyright © 1996 with permission from Elsevier Science.

Figure 2. The mechanism of an ATP-dependent DNA ligase reaction. Step 1, the amino group of the active site lysine attacks the -phosphate of ATP, where A represents adenosine, to form a phosphoamide-linked AMP with the elimination of PP_i. Step 2, the 5¢-phosphate at the nick attacks the activated phosphoryl group of the AMP to form an adenylated DNA. Step 3, the nonadenylated enzyme catalyses the attack of the 3¢-OH of the DNA at the nick upon the activated 5¢-phosphate to form the phosphodiester bond and release AMP. The reaction of an NAD⁺-dependent enzyme is analogous except that NAD⁺ provides the adenyl group and NMN is released rather than PP_i.

Figure 3. Motif alignments for DNA ligases and RNA capping enzymes. The ligases are grouped in the top section, capping enzymes in the bottom section. Enzymes are from human (Hu), vaccinia virus (vac), Desulfurolobus ambivalens (Dam), Methanococcus jannaschii (Mja), Saccharomyces cerevisiae (Sce), Schizosaccharomyces pombe (Spo), African swine fever virus (asf), mouse (Mus), and Chlorella virus (ChV). Interactions of conserved side-chains with moieties on the nucleotide (revealed by crystal structures of T7 ligase and Chlorella virus capping enzyme) are denoted by arrows at the bottom. The dashes before motif I indicate that N-terminal amino acids precede it. The numbers between hyphens indicate the number of amino acids between motifs.

Figure 4. Structure of bacteriophage T7 DNA ligase•ATP complex (Subramanya et al., 1996). The crystallographic coordinates for the structure are available from the Brookhaven Protein Data Bank (Accession Code 1A0I). A ribbon structure is shown for the protein and the ATP is coloured orange. The conserved motifs (see text and Figure 3) are coloured yellow (motif I), green (motif III), light blue (motif IIIa), red–orange (motif IV), magenta (motif V), and dark blue (motif VI).