ATP-dependent DNA ligases

Abstract

By catalyzing the joining of breaks in the phosphodiester backbone of duplex DNA, DNA ligases play a vital role in the diverse processes of DNA replication, recombination and repair. Three related classes of ATP-dependent DNA ligase are readily apparent in eukaryotic cells. Enzymes of each class comprise catalytic and non-catalytic domains together with additional domains of varying function. DNA ligase I is required for the ligation of Okazaki fragments during lagging-strand DNA synthesis, as well as for several DNA-repair pathways; these functions are mediated, at least in part, by interactions between DNA ligase I and the sliding-clamp protein PCNA. DNA ligase III, which is unique to vertebrates, functions both in the nucleus and in mitochondria. Two distinct isoforms of this enzyme, differing in their carboxy-terminal sequences, are produced by alternative splicing: DNA ligase IIIalpha has a carboxy-terminal BRCT domain that interacts with the mammalian DNA-repair factor XrccI, but both alpha and beta isoforms have an amino-terminal zinc-finger motif that appears to play a role in the recognition of DNA secondary structures that resemble intermediates in DNA metabolism. DNA ligase IV is required for DNA non-homologous end joining pathways, including recombination of the V(D)J immunoglobulin gene segments in cells of the mammalian immune system. DNA ligase IV forms a tight complex with Xrcc4 through an interaction motif located between a pair of carboxy-terminal BRCT domains in the ligase. Recent structural studies have shed light on the catalytic function of DNA ligases, as well as illuminating protein-protein interactions involving DNA ligases IIIalpha and IV.

Figure 1

Domain structures of ATP-dependent ligases. Schematic representation of the domain structures of DNA ligases I, IIIα, IIIβ and IV, together with ATP-dependent ligases from poxviruses (vaccinia, variola, fowlpox, and so on), the Chlorella virus of Paramecium bursaria PBCV-1, and archaea. Abbreviations: CD, catalytic domain; NCD, conserved non-catalytic domain; PBM, PCNA binding motif; NLS, nuclear localization signal; MTS, mitochondrial targeting sequence; ZnF, putative zinc finger; BRCT, BRCA carboxy-terminal-related domain. The red-boxed regions have had their structures solved crystallographically; the blue-boxed regions are found only in proteins targeted to mitochondria.

Figure 2

Structures of ATP-dependent DNA ligases. (a,b) Three-dimensional structures of (a) bacteriophage T7 DNA ligase complexed with ATP and (b) the Chlorella virus of Paramecium bursaria (PBCV-1) DNA ligase enzyme-adenylate complex, determined by X-ray crystallography. For the PBCV-1 enzyme, domains 1 and 2 (an OB fold) are indicated. (c,d) Structures of the BRCT domains from (c) the human DNA-repair factor Xrcc1 and (d) DNA ligaseIIIα, determined by X-ray crystallography and NMR (nuclear magnetic resonance), respectively. In each case, four short β strands form the core of the BRCT structure. The Xrcc1 core is flanked by three α helices (α1, α2 and α3) whereas that of DNA ligase IIIα is flanked by two only (α1 and α2). Theinteraction between the Xrcc1 and DNA ligase IIIα proteins in vivo is mediated by these BRCT domains. (e) Structure of a homodimer of Xrcc4 bound to a short peptide corresponding to amino acids 748-784 of human DNA ligase IV (shown in green). (f) Close-up view of the DNA ligase IV peptide bound to the helical tails of the Xrcc4 dimer. The peptide comprises a β hairpin followed by an α helix and lies asymmetrically across both Xrcc4 monomers. See text for details and references.