EMD-0388
Structure of bacteriophage T7 lagging-strand DNA polymerase (D5A/E7A)/Trx interacting with primase domains of two gp4 subunits (C and D), with gp4 helicase bound to a DNA fork and dTTP (LagL2)
EMD-0388
Single-particle7.1 Å
Deposition: 05/12/2018Map released: 06/03/2019
Last modified: 06/03/2019
Sample Organism:
Enterobacteria phage T7,
Escherichia coli
Sample: T7 DNA polymerase/Trx interacting with two primase domains (C and D) of T7 helicase (gp4) in complex with a fork DNA substrate and dTTP (from gp4-gp5 lagging-strand complex LagL2)
Deposition Authors: Gao Y, Fox T, Val N, Yang W
Sample: T7 DNA polymerase/Trx interacting with two primase domains (C and D) of T7 helicase (gp4) in complex with a fork DNA substrate and dTTP (from gp4-gp5 lagging-strand complex LagL2)
Deposition Authors: Gao Y, Fox T, Val N, Yang W
Structures and operating principles of the replisome.
Abstract:
Visualization in atomic detail of the replisome that performs concerted leading- and lagging-DNA strand synthesis at a replication fork has not been reported. Using bacteriophage T7 as a model system, we determined cryo-electron microscopy structures up to 3.2-angstroms resolution of helicase translocating along DNA and of helicase-polymerase-primase complexes engaging in synthesis of both DNA strands. Each domain of the spiral-shaped hexameric helicase translocates sequentially hand-over-hand along a single-stranded DNA coil, akin to the way AAA+ ATPases (adenosine triphosphatases) unfold peptides. Two lagging-strand polymerases are attached to the primase, ready for Okazaki fragment synthesis in tandem. A β hairpin from the leading-strand polymerase separates two parental DNA strands into a T-shaped fork, thus enabling the closely coupled helicase to advance perpendicular to the downstream DNA duplex. These structures reveal the molecular organization and operating principles of a replisome.
Visualization in atomic detail of the replisome that performs concerted leading- and lagging-DNA strand synthesis at a replication fork has not been reported. Using bacteriophage T7 as a model system, we determined cryo-electron microscopy structures up to 3.2-angstroms resolution of helicase translocating along DNA and of helicase-polymerase-primase complexes engaging in synthesis of both DNA strands. Each domain of the spiral-shaped hexameric helicase translocates sequentially hand-over-hand along a single-stranded DNA coil, akin to the way AAA+ ATPases (adenosine triphosphatases) unfold peptides. Two lagging-strand polymerases are attached to the primase, ready for Okazaki fragment synthesis in tandem. A β hairpin from the leading-strand polymerase separates two parental DNA strands into a T-shaped fork, thus enabling the closely coupled helicase to advance perpendicular to the downstream DNA duplex. These structures reveal the molecular organization and operating principles of a replisome.