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PDBsum entry 2cfm

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Ligase PDB id
2cfm
Jmol
Contents
Protein chain
561 a.a.
Ligands
AMP
Metals
_MG
Waters ×450

References listed in PDB file
Key reference
Title The closed structure of an archaeal DNA ligase from pyrococcus furiosus.
Authors H.Nishida, S.Kiyonari, Y.Ishino, K.Morikawa.
Ref. J Mol Biol, 2006, 360, 956-967. [DOI no: 10.1016/j.jmb.2006.05.062]
PubMed id 16820169
Abstract
DNA ligases join single-strand breaks in double-stranded DNA, and are essential to maintain genome integrity in DNA metabolism. Here, we report the 1.8 A resolution structure of Pyrococcus furiosus DNA ligase (PfuLig), which represents the first full-length atomic view of an ATP-dependent eukaryotic-type DNA ligase. The enzyme comprises the N-terminal DNA-binding domain, the middle adenylation domain, and the C-terminal OB-fold domain. The architecture of each domain resembles those of human DNA ligase I, but the domain arrangements differ strikingly between the two enzymes. The closed conformation of the two "catalytic core" domains at the carboxyl terminus in PfuLig creates a small compartment, which holds a non-covalently bound AMP molecule. This domain rearrangement results from the "domain-connecting" role of the helical extension conserved at the C termini in archaeal and eukaryotic DNA ligases. The DNA substrate in the human open-ligase is replaced by motif VI in the Pfu closed-ligase. Both the shapes and electrostatic distributions are similar between motif VI and the DNA substrate, suggesting that motif VI in the closed state mimics the incoming substrate DNA. Two basic residues (R531 and K534) in motif VI reside within the active site pocket and interact with the phosphate group of the bound AMP. The crystallographic and functional analyses of mutant enzymes revealed that these two residues within the RxDK sequence play essential and complementary roles in ATP processing. This sequence is also conserved exclusively among the covalent nucleotidyltransferases, even including mRNA-capping enzymes with similar helical extensions at the C termini.
Figure 1.
Figure 1. (a) Ribbon diagram illustrating the overall structure of P. furiosus DNA ligase (Pfu; colored blue), compared with that of human DNA ligase I (colored orange). The adenylation and OB-fold domains of Pfu DNA ligase are tightly closed, whereas those of human DNA ligase I exhibit an open conformation. The Figure was prepared using the program Chimera.^28 (b) Close-up view of the interface of the C-terminal helix and the adenylation domain. The contact surface is predominantly composed of hydrophilic interactions. Ball-and-stick models represent the amino acid residues involved in the interaction: basic residues are blue, acidic residues are red, and polar residues are purple.
Figure 3.
Figure 3. (a) Superimposed structures of PfuLig (blue) and hLigI (orange) around the AMP-binding region. The structures were overlapped manually by fitting the adenyl ring moiety of AMP, colored blue (Pfu) and orange (human). Residues in motif VI, except for RE(D)DK, are depicted as alanine. The bound DNA substrate in hLigI is depicted by a sheet representation and is colored pink. (b) Ribbon diagrams of the adenylation domains from PfuLig and hLigI, flanked by surface representations of motif VI (PfuLig) and the upstream region of the substrate DNA (hLigI), in which the colors of the atoms (nitrogen, blue; oxygen, red) are mapped onto the surface. Since the distribution patterns of the charged atoms on the surfaces of motif VI and the DNA are similar to each other, the replacement of motif VI with DNA may easily occur. Helix A3 of the adenylation domain is responsible for interacting with the acidic portions of both motif VI and DNA, so the C-terminal region of helix A3 should have an abundance of basic residues in the Pfu and human forms, but not in the ChV and T7 forms, implying that this feature is specific for the ligases that adopt an open-closed exchange mechanism with a C-terminal extension helix. (c) Superimposed structures of the RE(D)DK motifs from the closed (PfuLig, cyan) and the open (hLigI, orange) forms. This Figure was prepared by minimizing the distances between the corresponding atom pairs, except for the side-chains of D532 and E880.
The above figures are reprinted by permission from Elsevier: J Mol Biol (2006, 360, 956-967) copyright 2006.
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