Fork sensing and strand switching control antagonistic activities of RecQ helicases
Daniel Klaue,Daniela Kobbe,Felix E. Kemmerich,Alicja Kozikowska,Holger Puchta,Ralf Seidel,Ralf Seidel +6 more
TLDR
This work investigates the DNA unwinding of RecQ helicases from Arabidopsis thaliana, AtRECQ2 and AtRECZ3 at the single-molecule level using magnetic tweezers and provides a simple explanation for how different biological activities can be achieved by rather similar members of the RecQ family.Abstract:
RecQ helicases have essential roles in maintaining genome stability during replication and in controlling double-strand break repair by homologous recombination. Little is known about how the different RecQ helicases found in higher eukaryotes achieve their specialized and partially opposing functions. Here, we investigate the DNA unwinding of RecQ helicases from Arabidopsis thaliana, AtRECQ2 and AtRECQ3 at the single-molecule level using magnetic tweezers. Although AtRECQ2 predominantly unwinds forked DNA substrates in a highly repetitive fashion, AtRECQ3 prefers to rewind, that is, to close preopened DNA forks. For both enzymes, this process is controlled by frequent strand switches and active sensing of the unwinding fork. The relative extent of the strand switches towards unwinding or towards rewinding determines the predominant direction of the enzyme. Our results provide a simple explanation for how different biological activities can be achieved by rather similar members of the RecQ family.read more
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Camera-based three-dimensional real-time particle tracking at kHz rates and Ångström accuracy
Alexander Huhle,Daniel Klaue,Hergen Brutzer,Peter Daldrop,Sihwa Joo,Oliver Otto,Ulrich F. Keyser,Ralf Seidel,Ralf Seidel +8 more
TL;DR: This work demonstrates that camera-based imaging can provide a similar performance for all three dimensions of particle tracking with Ångström accuracy as laser detection through photodiodes, and provides a simple and robust way for high-resolution tweezers experiments using multiple particles at a time.
Journal ArticleDOI
Mechanistic insight into the interaction of BLM helicase with intra-strand G-quadruplex structures.
Sujoy Chatterjee,Jennifer Zagelbaum,Pavel Savitsky,Andreas Sturzenegger,Diana Huttner,Pavel Janscak,Ian D. Hickson,Opher Gileadi,Eli Rothenberg +8 more
TL;DR: It is shown that the activity of BLM is substrate dependent, and highly regulated by a short ssDNA segment that separates the G4 motif from dsDNA, and a model is presented that proposes a unique role for G4 structures in modulating theActivity of DNA processing enzymes.
Journal ArticleDOI
Human DNA2 possesses a cryptic DNA unwinding activity that functionally integrates with BLM or WRN helicases
TL;DR: It is shown that the helicase of hDNA2 functionally integrates with BLM or WRN helicases to promote dsDNA degradation by forming a heterodimeric molecular machine, which collectively suggests that the h DNA2 motor promotes the enzyme's capacity to degrade ds DNA in conjunction with BLMor WRN and thus promote the repair of broken DNA.
Journal ArticleDOI
Dynamic look at DNA unwinding by a replicative helicase
Seung Jae Lee,Salman Syed,Eric J. Enemark,Eric J. Enemark,Stephen Schuck,Arne Stenlund,Taekjip Ha,Leemor Joshua-Tor,Leemor Joshua-Tor,Leemor Joshua-Tor +9 more
TL;DR: The findings reveal that E1 employs a strand exclusion mechanism to unwind DNA with the N-terminal side leading at the replication fork, and DNA unwinding by E1 is modulated by the origin-recognition domain, suggesting a previously unsuspected role for this domain in regulating helicase activity.
Journal ArticleDOI
Extending the Range for Force Calibration in Magnetic Tweezers
Peter Daldrop,Hergen Brutzer,Alexander Huhle,Dominik J. Kauert,Ralf Seidel,Ralf Seidel,Ralf Seidel +6 more
TL;DR: The force calibration based on the long pendulum geometry will facilitate high-resolution magnetic-tweezers experiments that rely on short molecules and large forces, as well as highly parallelized measurements that use low frame rates.
References
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Journal ArticleDOI
Regulation of translocation polarity by helicase domain 1 in SF2B helicases.
TL;DR: Proteolytic DNA and mutational analysis has determined that SF2B helicases bind ssDNA with the same orientation as their 3′–5′ counterparts, and 5′–3′ translocation polarity requires conserved residues in HD1 and the FeS cluster containing domain.
Journal ArticleDOI
Single-stranded DNA Scanning and Deamination by APOBEC3G Cytidine Deaminase at Single Molecule Resolution
Gayan Senavirathne,Malgorzata Jaszczur,Paul A. Auerbach,Thomas G. Upton,Linda Chelico,Myron F. Goodman,David Rueda +6 more
TL;DR: The specific issue of deamination asymmetry within the general context of ssDNA scanning mechanisms is addressed and it is shown that Apo3G scanning trajectories, ssDNA contraction, and deamination efficiencies depend on motif sequence, location, and ionic strength.
Journal ArticleDOI
DnaB Helicase Activity Is Modulated by DNA Geometry and Force
TL;DR: In this article, the authors quantified translocation and unwinding by single DnaB molecules in three tethered DNA geometries held under tension, and found that the magnitude of this stimulation varies with the geometry of the tension applied to the DNA substrate, possibly due to interactions between the helicase and the occluded ssDNA strand.
Interaction between Arabidopsis Brca2 and its partners Rad51, Dmc1, and Dss1
TL;DR: The Arabidopsis orthologs of Brca2, a protein whose mutations are involved in breast cancer in humans, were previously shown to be essential at meiosis, and it is shown that both AtBrca2 proteins can interact with either AtRad51 or AtDmc1 in vitro, and that the N-terminal region of AtBRCa2 is responsible for these interactions.
Journal ArticleDOI
A prominent β-hairpin structure in the winged-helix domain of RECQ1 is required for DNA unwinding and oligomer formation
Bojana Lucic,Ying Zhang,Oliver N. King,Ramiro Mendoza-Maldonado,Matteo Berti,Frank H. Niesen,N.A. Burgess-Brown,Ashley C. W. Pike,Christopher D.O. Cooper,Opher Gileadi,Alessandro Vindigni +10 more
TL;DR: It is shown that the β-hairpin is required for the DNA unwinding and Holliday junction (HJ) resolution activity of full-length RECQ1, confirming that it represents an important determinant for the distinct substrate specificity of the five human RecQ helicases.