Asymmetric Regulation of Bipolar Single-stranded DNA Translocation by the Two Motors within Escherichia coli RecBCD Helicase

Xie, Fuqian; Wu, Colin G.; Weiland, Elizabeth; Lohman, Timothy M.

doi:10.1074/jbc.m112.423384

Cited by 24 publications

(84 citation statements)

References 42 publications

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“…Although it had been proposed that the motor activities of RecD and RecB are independent and uncoupled with RecD being the faster motor before chi recognition and RecB being the faster motor after chi recognition[11], our recent findings[15, 17, 18] suggest a more complex scenario. We found that RecBC (without RecD) possesses two distinct translocase activities that are controlled by the single RecB motor[15, 17].…”

Section: Introductionmentioning

confidence: 65%

“…As such, RecBC can move along two unpaired strands of ssDNA at the same rate in a concerted mechanism in which both translocases are tightly coupled to ATP hydrolysis by the RecB motor[15, 17]. This secondary translocase activity, that might represent a double stranded DNA translocase activity[15], also functions within RecBCD, hence the RecB and RecD motors are functionally, but asymmetrically, coupled due to the action of the secondary RecBC translocase[18]. That is, before chi, RecB regulates both 3’ to 5’ and 5’ to 3’ translocation, whereas RecD regulates only 5’ to 3’ translocation[18].…”

Section: Introductionmentioning

confidence: 99%

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Processive DNA Unwinding by RecBCD Helicase in the Absence of Canonical Motor Translocation

Simon

Sokoloski

Hao

et al. 2016

Journal of Molecular Biology

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Escherichia coli RecBCD is a DNA helicase/nuclease that functions in double-stranded DNA break repair. RecBCD possesses two motors (RecB, a 3′ to 5′ translocase, and RecD, a 5′ to 3′ translocase). Current DNA unwinding models propose that motor translocation is tightly coupled to base pair (bp) melting. However, some biochemical evidence suggests that DNA melting of multiple bp may occur separately from single stranded DNA translocation. To test this hypothesis, we designed DNA substrates containing reverse backbone polarity (RP) linkages that prevent ssDNA translocation of the canonical RecB and RecD motors. Surprisingly, we find that RecBCD can processively unwind DNA for at least 80 bp beyond the RP linkages. This ability requires an ATPase active RecB motor, the RecB “arm” domain and also the RecB nuclease domain, but not its nuclease activity. These results indicate that RecBCD can unwind duplex DNA processively in the absence of ssDNA translocation by the canonical motors and that the nuclease domain regulates the helicase activity of RecBCD.

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Section: Introductionmentioning

confidence: 65%

Section: Introductionmentioning

confidence: 99%

Processive DNA Unwinding by RecBCD Helicase in the Absence of Canonical Motor Translocation

Simon

Sokoloski

Hao

et al. 2016

Journal of Molecular Biology

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“…Although RecBCD is most often characterized using linear, double-stranded substrates, the enzyme will also unwind and nick cruciform DNA at its junction, and can unwind duplex DNA containing gaps on either strand [90, 91] (Figure 5D). There is also evidence to suggest that the complex may also have other important protein partners, including Polymerase I, and SbcC, another protein involved in the completion of replication that is homologous to human Mre11 [22, 92–94].…”

Section: Recbcd Is More Than a Helicase And Nucleasementioning

confidence: 99%

“…If [Mg +2 ]<[ATP], RecBCD unwinds DNA, nicking the strand at a Chi sequence, then continues to unwind the molecule [11, 12] D) In vitro substrates of RecBCD. In addition to unwinding double stranded DNA (i), the complex can translocate through single-stranded gaps on either strand (ii), and will unwind cruciform structures, making an incision (*) at the branch point (iii) [90, 91]. …”

Section: Figurementioning

confidence: 99%

RecBCD is required to complete chromosomal replication: Implications for double-strand break frequencies and repair mechanisms

et al. 2015

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Several aspects of the mechanism of homologous double strand break repair remain unclear. Although intensive efforts have focused on how recombination reactions initiate, far less is known about the molecular events that follow. Based upon biochemical studies, current models propose that RecBCD processes double strand ends and loads RecA to initiate recombinational repair. However, recent studies have shown that RecBCD plays a critical role in completing replication events on the chromosome through a mechanism that does not involve RecA or recombination. Here, we examine several studies, both early and recent, that suggest RecBCD also operates late in the recombination process- after initiation, strand invasion, and crossover resolution have occurred. Similar to its role in completing replication, we propose a model in which RecBCD is required to resect and resolve the DNA synthesis associated with homologous recombination at the point where the missing sequences on the broken molecule have been restored. We explain how the impaired ability to complete chromosome replication in recBC and recD mutants is likely to account for the loss of viability and genome instability in these mutants, and conclude that spontaneous double strand breaks and replication fork collapse occur far less frequently than previously speculated.

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“…Interestingly, the presence of these two different motors allows RecBCD to translocate along ssDNA either from 3′ to 5′ or from 5′ to 3′ (59) (see Figure 3). The rate of RecBCD translocation in the 5′-3′ direction along ssDNA is faster than the rate of translocation in the 3′-5′ direction (60); at saturating ATP concentrations this rate is (1515 ± 33) nt/s for 3′-5′ translocation and (2037 ± 109) nt/s for 5′-3′ translocation. There is no difference in the kinetic step-size for these two directions of RecBCD translocation and both are ∼4 nucleotides (60).…”

Section: Sf1 Family Helicasesmentioning

confidence: 96%

All motors have to decide is what to do with the DNA that is given them

Briggs

Fischer

2014

Biomolecular Concepts

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DNA translocases are a diverse group of molecular motors responsible for a wide variety of cellular functions. The goal of this review is to identify common aspects in the mechanisms for how these enzymes couple the binding and hydrolysis of ATP to their movement along DNA. Not surprisingly, the shared structural components contained within the catalytic domains of several of these motors appear to give rise to common aspects of DNA translocation. Perhaps more interesting, however, are the differences between the families of translocases and the potential associated implications both for the functions of the members of these families and for the evolution of these families. However, as there are few translocases for which complete characterizations of the mechanisms of DNA binding, DNA translocation, and DNA-stimulated ATPase have been completed, it is difficult to form many inferences. We therefore hope that this review motivates the necessary further experimentation required for broader comparisons and conclusions.

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Asymmetric Regulation of Bipolar Single-stranded DNA Translocation by the Two Motors within Escherichia coli RecBCD Helicase

Cited by 24 publications

References 42 publications

Processive DNA Unwinding by RecBCD Helicase in the Absence of Canonical Motor Translocation

Processive DNA Unwinding by RecBCD Helicase in the Absence of Canonical Motor Translocation

RecBCD is required to complete chromosomal replication: Implications for double-strand break frequencies and repair mechanisms

All motors have to decide is what to do with the DNA that is given them

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