Nucleic Acids Research Advance Access published April 21, 2015 Nucleic Acids Research, 2015 1 doi: 10.1093/nar/gkv373 Crystal structure of the Bloom’s syndrome helicase indicates a role for the HRDC domain in conformational changes Joseph A. Newman1,†, Pavel Savitsky1,†, Charles K. Allerston1, Anna H. Bizard2, Ozg¨ un¨ Ozer¨ 2, Kata Sarlos´ 2, Ying Liu2, Els Pardon3,4, Jan Steyaert3,4, Ian D. Hickson2 and Opher Gileadi1,*
1Structural Genomics Consortium, University of Oxford, ORCRB, Roosevelt Drive, Oxford OX3 7DQ, UK, 2Center for Chromosome Stability and Center for Healthy Aging, Department of Cellular and Molecular Medicine, University of Copenhagen, Panum Institute, Building 18.1, Blegdamsvej 3B, 2200 Copenhagen N, Denmark, 3Structural Biology Downloaded from Brussels, Vrije Universiteit Brussel, Pleinlaan 2 , 1050 Brussels, Belgium and 4Structural Biology Research Center, VIB, Brussel, Pleinlaan 2, 1050 Brussels, Belgium
Received January 13, 2015; Revised March 27, 2015; Accepted April 03, 2015 http://nar.oxfordjournals.org/
ABSTRACT some instability, including chromatid gaps and breaks (4), various chromosome structural rearrangements (5)andan Bloom’s syndrome helicase (BLM) is a member of the increase in the number of sister chromatid exchange (SCE) RecQ family of DNA helicases, which play key roles in events, the latter serving as a distinguishing feature for the the maintenance of genome integrity in all organism clinical diagnosis of BS (6). groups. We describe crystal structures of the BLM Bloom’s syndrome helicase (BLM), like all RecQ-family helicase domain in complex with DNA and with an an- helicases, acts as a 3 to 5 helicase (7) on a wide variety tibody fragment, as well as SAXS and domain asso- of DNA substrates including forked duplexes, G quadru- at Chadwick & RAL Libraries on April 23, 2015 ciation studies in solution. We show an unexpected plexes, 4-way junctions (8) and displacement loops (D nucleotide-dependent interaction of the core heli- loops) (9). It forms a multi protein complex with hu- ␣ case domain with the conserved, poorly character- man topoisomerase III (10), RMI1 (11) and RMI2 (12), ized HRDC domain. The BLM–DNA complex shows termed the ‘dissolvasome’, which can promote the con- vergent branch migration (13) and decatenation of dou- an unusual base-flipping mechanism with unique ble Holliday junction intermediates formed during homolo- positioning of the DNA duplex relative to the heli- gous recombination. This dissolution reaction prevents the case core domains. Comparison with other crystal exchange of genetic material flanking two homologous se- structures of RecQ helicases permits the definition quences engaged in homologous recombination (14,15). of structural transitions underlying ATP-driven heli- The BLM protein is a 1417 amino acid, 159-kDa case action, and the identification of a nucleotide- polypeptide with multiple structural domains (Figure 1A). regulated tunnel that may play a role in interactions At its centre (residues 639–1290) is a catalytic core con- with complex DNA substrates. served amongst RecQ-family helicases. A large N-terminal domain (residues 1–638) is believed to play a role in regu- lation and oligomerization of BLM (16,17), and is impor- INTRODUCTION tant in mediating interactions with partner proteins (10,18– Bloom’s syndrome (BS) is an extremely rare autosomal re- 19). The C-terminal region (residues 1291–1417), also in- cessive disorder that is characterized by shortness of stature, volved in protein interactions (10), is believed to be pre- a distinctive skin rash and the predisposition to the devel- dominantly unstructured, and contains a nuclear localiza- opment of a wide spectrum of cancers at an early age (1). tion signal (20). We have focussed this study on the core do- Individuals with BS carry mutations in BLM that result ei- main, which is responsible for the basic functions of DNA ther in expression of truncated proteins or alterations of key unwinding. conserved residues within the RecQ helicase domain (1–3). The core domain itself is composed of subdomains, with On a cellular level, BS is characterized by excessive chromo- varying degrees of evolutionary conservation (Figure 1A).
*To whom correspondence should be addressed. Tel: +44 1865 617572; Fax: +44 1865 617575; Email: [email protected] †These authors contributed equally to the paper as first authors. Present address: Charles K. Allerston, Dart Neuroscience LLC San Diego, CA 92121, USA.