The Escherichia coli dnaB replication protein is a DNA helicase.

LeBowitz, Jonathan H.; McMacken, Roger

doi:10.1016/s0021-9258(17)38564-2

Cited by 485 publications

(133 citation statements)

References 44 publications

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“…This difference implies that dSaCas9 could act as a more effective DNA roadblock compared to dSpCas9 to perturb DNA‐based transactions, such as DNA replication. To test this hypothesis, we first chose to use DnaB (in complex with DnaC, hereafter referred to as DnaB), which is an E. coli replicative helicase that catalyzes the separation of dsDNA during DNA replication (LeBowitz & McMacken, ), to mimic the replication fork and examine the outcome of its encountering with a DNA‐bound dSaCas9. A previously developed single‐molecule optical tweezer assay was employed (Sun et al , , , ).…”

Section: Resultsmentioning

confidence: 99%

Dynamics of Staphylococcus aureus Cas9 in DNA target Association and Dissociation

Zhang

Hou

et al. 2020

EMBO Reports

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Staphylococcus aureus Cas9 (SaCas9) is an RNA-guided endonuclease that targets complementary DNA adjacent to a protospacer adjacent motif (PAM) for cleavage. Its small size facilitates in vivo delivery for genome editing in various organisms. Herein, using single-molecule and ensemble approaches, we systemically study the mechanism of SaCas9 underlying its interplay with DNA. We find that the DNA binding and cleavage of SaCas9 require complementarities of 6and 18-bp of PAM-proximal DNA with guide RNA, respectively. These activities are mediated by two steady interactions among the ternary complex, one of which is located approximately 6 bp from the PAM and beyond the apparent footprint of SaCas9 on DNA. Notably, the other interaction within the protospacer is significantly strong and thus poses DNA-bound SaCas9 a persistent block to DNA-tracking motors. Intriguingly, after cleavage, SaCas9 autonomously releases the PAM-distal DNA while retaining binding to the PAM. This partial DNA release immediately abolishes its strong interaction with the protospacer DNA and consequently promotes its subsequent dissociation from the PAM. Overall, these data provide a dynamic understanding of SaCas9 and instruct its effective applications.

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Section: Resultsmentioning

confidence: 99%

Dynamics of Staphylococcus aureus Cas9 in DNA target Association and Dissociation

Zhang

Hou

et al. 2020

EMBO Reports

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“…DNA helicase plays an important role in DNA replication by unwinding double-stranded DNA before DNA polymerase synthesizes the daughter DNA strand (LeBowitz & McMacken, 1986). The p143 gene of AcMNPV has helicase activity (McDougal & Guarino, 2001) and has been implicated as a host-range factor (Croizier et al, 1994;Kamita & Maeda, 1997).…”

Section: Discussionmentioning

confidence: 99%

Slow cell infection, inefficient primary infection and inability to replicate in the fat body determine the host range of Thysanoplusia orichalcea nucleopolyhedrovirus

Wang

Salem

Lynn

et al. 2008

Journal of General Virology

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Thysanoplusia orichacea multicapsid nucleopolyhedrovirus (ThorMNPV) carrying an enhanced green fluorescent protein (EGFP) gene expression cassette (vThGFP) was used to study host-range mechanisms. Infection kinetics showed that vThGFP replication in Sf21 cells was too slow to suppress cell growth. Wide-host-range Autographa californica MNPV (AcMNPV) could speed up vThGFP infection and enhance the vThGFP infection rate in Sf21 cells. The enhancement was not due to recombination, as no recombinant virus was isolated from coinfection by plaque assay. No improvement of vThGFP infection in Sf21 was found by AcMNPV cosmid transactivation assay. However, culture medium from Sf21 cells infected with AcMNPV did enhance vThGFP replication in Sf21. Third-instar larvae of Spodoptera frugiperda, S. exigua and Helicoverpa zea were not killed by feeding with vThGFP polyhedra but were killed by intrahaemocoelic injection using budded viruses (BVs). This suggested that insufficient BVs were generated during the primary infection in the midgut. vThGFP infected haemocytes, tracheae and Malpighian tubules but not fat bodies of larvae of S. frugiperda, S. exigua and H. zea. Third-instar S. frugiperda larvae co-infected by injection with vThGFP and vAcDsRed2, an AcMNPV expressing a red fluorescent protein gene, showed EGFP expression in the fat body. This result suggests that vAcDsRed2 could help vThGFP to replicate in the fat body or trans-activate EGFP expression in the fat body. All these results suggested that slow cell infection, insufficient primary infection and inability to replicate in the fat body control the host range of ThorMNPV.

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“…In bacteria, the toroid hexameric helicases that drive bidirectional DNA replication from the chromosome origin are loaded around ssDNA ( 3 ). To permit helicase ssDNA access, bacterial chromosome origins are opened by the ubiquitous initiation protein, DnaA ( 4 ).…”

Section: Introductionmentioning

confidence: 99%

Evidence for a chromosome origin unwinding system broadly conserved in bacteria

Pelliciari

Dong

Gao

et al. 2021

Nucleic Acids Research

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Genome replication is a fundamental requirement for the proliferation of all cells. Throughout the domains of life, conserved DNA replication initiation proteins assemble at specific chromosomal loci termed replication origins and direct loading of replicative helicases (1). Despite decades of study on bacterial replication, the diversity of bacterial chromosome origin architecture has confounded the search for molecular mechanisms directing the initiation process. Recently a basal system for opening a bacterial chromosome origin (oriC) was proposed (2). In the model organism Bacillus subtilis, a pair of double-stranded DNA (dsDNA) binding sites (DnaA-boxes) guide the replication initiator DnaA onto adjacent single-stranded DNA (ssDNA) binding motifs (DnaA-trios) where the protein assembles into an oligomer that stretches DNA to promote origin unwinding. We report here that these core elements are predicted to be present in the majority of bacterial chromosome origins. Moreover, we find that the principle activities of the origin unwinding system are conserved in vitro and in vivo. The results suggest that this basal mechanism for oriC unwinding is broadly functionally conserved and therefore may represent an ancestral system to open bacterial chromosome origins.

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The Escherichia coli dnaB replication protein is a DNA helicase.

Cited by 485 publications

References 44 publications

Dynamics of Staphylococcus aureus Cas9 in DNA target Association and Dissociation

Dynamics of Staphylococcus aureus Cas9 in DNA target Association and Dissociation

Slow cell infection, inefficient primary infection and inability to replicate in the fat body determine the host range of Thysanoplusia orichalcea nucleopolyhedrovirus

Evidence for a chromosome origin unwinding system broadly conserved in bacteria

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