DNA helicase and helicase-nuclease enzymes with a conserved iron-sulfur cluster

Abstract

Conserved Iron-Sulfur (Fe-S) clusters are found in a growing family of metalloproteins that are implicated in prokaryotic and eukaryotic DNA replication and repair. Among these are DNA helicase and helicase-nuclease enzymes that preserve chromosomal stability and are genetically linked to diseases characterized by DNA repair defects and/or a poor response to replication stress. Insight to the structural and functional importance of the conserved Fe-S domain in DNA helicases has been gleaned from structural studies of the purified proteins and characterization of Fe-S cluster site-directed mutants. In this review, we will provide a current perspective of what is known about the Fe-S cluster helicases, with an emphasis on how the conserved redox active domain may facilitate mechanistic aspects of helicase function. We will discuss testable models for how the conserved Fe-S cluster might operate in helicase and helicase-nuclease enzymes to conduct their specialized functions that help to preserve the integrity of the genome.

Figure 1.

Fe–S cluster in the helicase domain of XPD/FANCJ family of DNA helicases. (A) Relative positions of the conserved Fe–S domain (yellow) are shown for the SF2 XPD/FANCJ family of DNA helicases. Homo sapiens (Hs) FANCJ, XPD, ChlR1, RTEL, Ferroplasma acidarmanus (Fac) XPD, Saccharomyces cerevisiae (Sc) Rad3 and E. coli (Ec) DinG are shown. Helicase domain is shown in purple. The BRCA1-binding domain of HsFANCJ is shown in green. The PCNA interaction motif (PIP) of HsRTEL is shown in blue. (B) Sequence alignment of the Fe–S domain helicases. The four conserved cysteine residues are highlighted in orange. The XPD amino acid substitution R112H genetically linked to trichothiodystrophy is indicated. The FANCJ amino acid substitutions M299I associated with breast cancer and A349P linked to Fanconi Anemia are shown. Amino acid positions corresponding to the beginning and end of the Fe–S cluster are indicated. Amino acids residing between conserved cysteines may not be shown due to space limitations.

Figure 2.

Structure of the apo-XPD catalytic core from Sulfolobus acidocaldarius. (A) The four XPD catalytic core domains are depicted in boxes for helicase domain (HD) 1 (cyan), HD2 (green), 4FeS (orange) with cysteine (C) residues indicated and Arch (purple) domains. Conserved helicase motifs (red bars with white labels) are shown. (B) XPD catalytic core fold and domains (ribbons). HD1 (cyan) and HD2 (green) form the nucleotide-binding pocket. Front view (left) shows that an arch is formed by the insertion of 4FeS (orange) and Arch (purple) domains, into HD1. Side view (right) shows that HD2 protrudes from the flat box formed by HD1, 4FeS and Arch as well as the HD2 helix-loop-helix insertion (green). Figure was provided by Drs Jill Fuss and John Tainer (45).

Figure 3.

Structure of the T. acidophilium XPD–DNA complex. Overall structure of XPD, shown in a transparent surface representation, with the two RecA-like domains in yellow and red, the FeS cluster domain in cyan, and the arch domain in green. The 4Fe–4S cluster is shown by the spheres with orange (Fe atom) and yellow (Cys residue) colors. The DNA identified in the electron density is shown in orange. Combination of experimentally verified DNA is shown in orange with modeled DNA shown in gray. Figure was provided by Drs Jochen Kuper and Caroline Kisker (45).

Figure 4.

Model depicting the coordinate action of an accessory Fe–S cluster 5′–3′ helicase with the MCM 3′–5′ helicase to clear a protein blockade encountered by the eukaryotic MCM/ replication machinery. For simplicity, the replication machinery is not shown. Model is adapted by analogy from one proposed for E. coli DNA replication (90).

Figure 5.

Fe–S cluster in the nuclease domain of eukaryotic Dna2 and AddB of the bacterial AddAB helicase–nuclease. Positions of the four conserved cysteine residues in the nuclease domain (green) are shown for human Dna2 and Bacillus subtilis AddB (102). The helicase domain is shown in purple. For Dna2, the nuclease and helicase domains reside in the same polypeptide. AddAB exists as a protein dimer with the Fe–S nuclease domain residing in AddB. AddA contains a second nuclease domain as well as a helicase domain.

Figure 6.

Models depicting functional cooperativity between Fe–S cluster helicase molecules as they unwind DNA duplexes (A), or remove proteins bound to double-stranded DNA (B) or single-stranded DNA (C). Models are adapted by analogy from ones proposed for bacteriophage T4 Dda helicase (118,120).