RecQ helicases: multiple structures for multiple functions?

Abstract

Approximately 1% of the open reading frames in the human genome encode proteins that function as DNA or RNA helicases. These enzymes act in all aspects of nucleic acid metabolism where the complementary strands of DNA:DNA or DNA:RNA duplexes require to be transiently opened. However, they perform wider roles in nucleic acid metabolism due to their ability to couple the energy derived from hydrolysis of ATP to their unidirectional translocation along strands of DNARNA. In this way, helicases can displace proteins from DNARNA, drive the migration of DNA junctions (such as the Holliday junction recombination intermediate), or generate superhelical tension in nucleic acid duplexes. Here, we review a subgroup of DNA helicase enzymes, the RecQ family, that has attracted considerable interest in recent years due to their role not only in suppression of genome instability, but also in the avoidance of human disease. We focus particularly on the protein structural motifs and the multiple assembly states that characterize RecQ helicases and discuss novel biophysical techniques to study the different RecQ structures present in solution. We also speculate on the roles of the different domains and oligomeric forms in defining which DNA structures will represent substrates for RecQ helicase-mediated transactions.

Figure 1. Schematic of selected members of the RecQ family of DNA helicases.

The respective organism is shown on the left, and the size (in amino acids) and name of each protein are indicated on the right. All proteins are aligned according to the conserved helicase domain, which is shown in yellow. The conserved RQC and HRDC domains are shown in red and green, respectively. The exonuclease domain in the amino-terminal region of WRN and its orthologs is shown in pink. Regions containing patches of acidic residues are shown in blue. The nuclear localization signal sequences identified at the extreme carboxyl terminus of certain family members is shown as a brown bar. The remaining portions of each protein (gray) represent regions that are poorly conserved. The sizes of the individual domains are not to scale. At least three splice variants of the human RECQ5 protein are expressed, of which only the largest (β isoform) is shown.

Figure 2. Schematic of the structural and functional properties of the RQC domain.

The RQC domain is divided into two subdomains, the Zn²⁺-binding domain (yellow) and the WH domain (green). The ribbon representations of a Zn²⁺ and WH domain of human RECQ1 are shown. The four Cys residues of the Zn²⁺ domain that coordinate the Zn ion (light blue sphere) are indicated. The Tyr residue (Y564) at the tip of the hairpin loop of the WH domain of RECQ1 is highlighted, and the DNA recognition helix is shown in blue. Putative functional properties of the Zn²⁺ and WH domains are indicated below.

Figure 3. Schematic of the structural and functional properties of the HRDC domain.

The structure of the HRDC domain of WRN is shown. Putative functional properties of HRDC domain are indicated below.

Figure 4. The human RECQ1 helicase exists in different oligomeric forms.

(A) Chromatographic profiles of the human RECQ1 in the absence (left) and presence (right) of ATPγS eluting from a Superdex200 HR 10∕30 gel filtration column. The protein species were detected by protein fluorescence (λ_excitation=290 nm and λ_emission=340 nm). Approximately 40 μg of recombinant RECQ1 were loaded at a final concentration of 1 μM. The protein elutes in two main peaks. The first peak (PK1) corresponds to a calculated molecular mass of approximately 400 kDa, whereas the second peak (PK2) corresponds to a calculated molecular mass of approximately 155 kDa. (Note that ATPγS shifts the equilibrium toward PK2). (B) Schematic of the two quaternary forms of RECQ1: higher-order oligomers (probably tetramers) exhibit DNA strand annealing as well as four-way Holliday junction resolution activity, while the lower-oligomers, probably dimers, are required for DNA unwinding. The surface rendering of 3D EM reconstructions of the higher RECQ1 assembly state were obtained as previously described (Muzzolini et al., 2007). The ribbon representation of RECQ1 was generously provided by Opher Gileadi from the Structural Genomics Consortium (SGC), Oxford, UK. The structure of the Mg²⁺-ADP bound RECQ1 molecule (PDB id: 2V1X) shows the two RecA-like domains (colored in red and blue), the Zn²⁺-binding domain (yellow), and the WH domain (green).