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The ATPase of the phi29 DNA packaging motor is a member of the hexameric AAA+ superfamily.

ABSTRACT: The AAA+ superfamily of proteins is a class of motor ATPases performing a wide range of functions that typically exist as hexamers. The ATPase of phi29 DNA packaging motor has long been a subject of debate in terms of stoichiometry and mechanism of action. Here, we confirmed the stoichiometry of phi29 motor ATPase to be a hexamer and provide data suggesting that the phi29 motor ATPase is a member of the classical hexameric AAA+ superfamily. Native PAGE, EMSA, capillary electrophoresis, ATP titration, and binomial distribution assay show that the ATPase is a hexamer. Mutations in the known Walker motifs of the ATPase validated our previous assumptions that the protein exists as another member of this AAA+ superfamily. Our data also supports the finding that the phi29 DNA packaging motor uses a revolution mechanism without rotation or coiling (Schwartz et al., this issue).

SUBMITTER: Schwartz C

PROVIDER: S-EPMC3700617 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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The ATPase of the phi29 DNA packaging motor is a member of the hexameric AAA+ superfamily.

Schwartz Chad C De Donatis Gian Marco GM Fang Huaming H Guo Peixuan P

Virology 20130522 1

The AAA+ superfamily of proteins is a class of motor ATPases performing a wide range of functions that typically exist as hexamers. The ATPase of phi29 DNA packaging motor has long been a subject of debate in terms of stoichiometry and mechanism of action. Here, we confirmed the stoichiometry of phi29 motor ATPase to be a hexamer and provide data suggesting that the phi29 motor ATPase is a member of the classical hexameric AAA+ superfamily. Native PAGE, EMSA, capillary electrophoresis, ATP titra ...[more]

PMID: 23706809

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Revolution rather than rotation of AAA+ hexameric phi29 nanomotor for viral dsDNA packaging without coiling.

Project description:It has long been believed that the DNA-packaging motor of dsDNA viruses utilizes a rotation mechanism. Here we report a revolution rather than rotation mechanism for the bacteriophage phi29 DNA packaging motor. The phi29 motor contains six copies of the ATPase (Schwartz et al., this issue); ATP binding to one ATPase subunit stimulates the ATPase to adopt a conformation with a high affinity for dsDNA. ATP hydrolysis induces a new conformation with a lower affinity, thus transferring the dsDNA to an adjacent subunit by a power stroke. DNA revolves unidirectionally along the hexameric channel wall of the ATPase, but neither the dsDNA nor the ATPase itself rotates along its own axis. One ATP is hydrolyzed in each transitional step, and six ATPs are consumed for one helical turn of 360°. Transition of the same dsDNA chain along the channel wall, but at a location 60° different from the last contact, urges dsDNA to move forward 1.75 base pairs each step (10.5bp per turn/6ATP=1.75bp per ATP). Each connector subunit tilts with a left-handed orientation at a 30° angle in relation to its vertical axis that runs anti-parallel to the right-handed dsDNA helix, facilitating the one-way traffic of dsDNA. The connector channel has been shown to cause four steps of transition due to four positively charged lysine rings that make direct contact with the negatively charged DNA phosphate backbone. Translocation of dsDNA into the procapsid by revolution avoids the difficulties during rotation that are associated with DNA supercoiling. Since the revolution mechanism can apply to any stoichiometry, this motor mechanism might reconcile the stoichiometry discrepancy in many phage systems where the ATPase has been found as a tetramer, hexamer, or nonamer.

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Structure of the bacteriophage phi29 DNA packaging motor.

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ATP/ADP modulates gp16-pRNA conformational change in the Phi29 DNA packaging motor.

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Fingerprinting of Peptides with a Large Channel of Bacteriophage Phi29 DNA Packaging Motor.

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