Insights into Strand Displacement and Processivity from the Crystal Structure of the Protein-Primed DNA Polymerase of Bacteriophage φ29

Kamtekar, S.; Berman, Andrea J.; Wang, Jimin; Lázaro, José Portolés; Vega, Miguel de; Blanco, Luis; Salas, Margarita; Steitz, Thomas A.

doi:10.1016/j.molcel.2004.12.006

Cited by 37 publications

(78 citation statements)

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“…Improvement of the heavy-atom phases by solvent-flipping and cross-crystal averaging, together with B-factor sharpening of the amplitudes, provided maps into which the main chain could be positioned. The sequence register of polymerase in these maps could be established based on previous high-resolution structures of polymerase (Kamtekar et al, 2004), but the sequence for terminal protein could not be assigned at this resolution.…”

Section: Structure Determinationmentioning

confidence: 99%

“…The structure of the polymerase:terminal protein heterodimer The structure of f29 DNA polymerase, as previously described, has the architecture of a canonical B-family DNA polymerase with two additional subdomains unique to protein-primed polymerases (Kamtekar et al, 2004). It contains Figure 1 Electron density for a helix of terminal protein near the active site of polymerase.…”

Section: Structure Determinationmentioning

confidence: 99%

“…The catalytic palm subdomain of f29 DNA polymerase can be aligned with the palm of the B-family DNA polymerase from bacteriophage RB69 (Franklin et al, 2001;Kamtekar et al, 2004). As described previously, this alignment allows placement of the primer, template, and incoming nucleotide from a ternary complex of RB69 polymerase onto f29 DNA polymerase (Kamtekar et al, 2004). The spatial overlap of the priming domain of terminal protein with modeled primer:template as well as its negative charge and overall dimensions suggest that it mimics DNA in its interactions with polymerase ( Figure 3).…”

Section: Structure Determinationmentioning

confidence: 99%

“…Two sequence insertions, TPR1 and TPR2 (Blasco et al, 1990;Dufour et al, 2000), found only in the polymerase domains of protein-primed DNA polymerases, formed discrete subdomains in the structure (Kamtekar et al, 2004). While TPR1 abutted the polymerase domain in a position consistent with interaction with terminal protein and DNA, TPR2 appeared to contribute to both efficient strand displacement and processivity.…”

Section: Introductionmentioning

confidence: 99%

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The φ29 DNA polymerase:protein-primer structure suggests a model for the initiation to elongation transition

Kamtekar

Berman

Wang

et al. 2006

EMBO J

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160

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The absolute requirement for primers in the initiation of DNA synthesis poses a problem for replicating the ends of linear chromosomes. The DNA polymerase of bacteriophage /29 solves this problem by using a serine hydroxyl of terminal protein to prime replication. The 3.0 Å resolution structure shows one domain of terminal protein making no interactions, a second binding the polymerase and a third domain containing the priming serine occupying the same binding cleft in the polymerase as duplex DNA does during elongation. Thus, the progressively elongating DNA duplex product must displace this priming domain. Further, this heterodimer of polymerase and terminal protein cannot accommodate upstream template DNA, thereby explaining its specificity for initiating DNA synthesis only at the ends of the bacteriophage genome. We propose a model for the transition from the initiation to the elongation phases in which the priming domain of terminal protein moves out of the active site as polymerase elongates the primer strand. The model indicates that terminal protein should dissociate from polymerase after the incorporation of approximately six nucleotides.

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Section: Structure Determinationmentioning

confidence: 99%

Section: Structure Determinationmentioning

confidence: 99%

Section: Structure Determinationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

The φ29 DNA polymerase:protein-primer structure suggests a model for the initiation to elongation transition

Kamtekar

Berman

Wang

et al. 2006

EMBO J

Self Cite

160

View full text Add to dashboard Cite

show abstract

“…We use φ29, a polymerase known for its exceptional strand displacement activity, to push a DNA cargo. Researchers have studied the structure of φ29 polymerase and have provided useful insights into its exceptional strand displacement and processivity, and have deduced its translocation mechanism [9,16,17,32].…”

Section: Our Contributionmentioning

confidence: 99%

A DNA Nanotransport Device Powered by Polymerase ϕ29

2008

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Abstract. Polymerases are a family of enzymes responsible for copying or replication of nucleic acids (DNA or RNA) templates and hence sustenance of life processes. In this paper, we present a method to exploit a strand-displacing polymerase φ29 as a driving force for nanoscale transportation devices. The principle idea behind the device is strong strand displacement ability of φ29, which can displace any DNA strand from its template while extending a primer hybridized to the template. This capability of φ29 is used to power the movement of a target nanostructure on a DNA track. The major advantage of using a polymerase driven nanotransportation device as compared to other existing nanorobotical devices is its speed. φ29 polymerase can travel at the rate of 2000 nucleotides per minute [1] at room temperature, which translates to approximately 680 nanometers per minute on a nanostructure. We also demonstrate transportation of a DNA cargo on a DNA track with the help of fluorescence resonance electron transfer (FRET) data.

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