title: The dynamic pause-unpackaging state, an off-translocation recovery state of a DNA packaging motor from bacteriophage T4.
abstract: Tailed bacteriophages and herpes viruses use powerful ATP-driven molecular motors to translocate their viral genomes into a preformed capsid shell. The bacteriophage T4 motor, a pentamer of the large terminase protein (gp17) assembled at the portal vertex of the prohead, is the fastest and most powerful known, consistent with the need to package a ~170-kb viral genome in approximately 5 min. Although much is known about the mechanism of DNA translocation, very little is known about how ATP modulates motor-DNA interactions. Here, we report single-molecule measurements of the phage T4 gp17 motor by using dual-trap optical tweezers under different conditions of perturbation. Unexpectedly, the motor pauses randomly when ATP is limiting, for an average of 1 s, and then resumes translocation. During pausing, DNA is unpackaged, a phenomenon so far observed only in T4, where some of the packaged DNA is slowly released. We propose that the motor pauses whenever it encounters a subunit in the apo state with the DNA bound weakly and incorrectly. Pausing allows the subunit to capture ATP, whereas unpackaging allows scanning of DNA until a correct registry is established. Thus, the "pause-unpackaging" state is an off-translocation recovery state wherein the motor, sometimes by taking a few steps backward, can bypass the impediments encountered along the translocation path. These results lead to a four-state mechanochemical model that provides insights into the mechanisms of translocation of an intricately branched concatemeric viral genome.
authors.affiliation: Department of Biology, The Catholic University of America, Washington, DC 20064, USA.
authors.forename: Vishal I
authors.initials: VI
authors.lastname: Kottadiel
authors.forename: Venigalla B
authors.initials: VB
authors.lastname: Rao
authors.forename: Yann R
authors.initials: YR
authors.lastname: Chemla
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D017122:Bacteriophage T4 / Q000502:physiology*; D042003:DNA Packaging / Q000502:physiology*; D007700:Kinetics; D008954:Models, Biological; D056004:Molecular Dynamics Simulation; D020409:Molecular Motor Proteins / Q000378:metabolism*; D052898:Optical Tweezers; D014764:Viral Proteins / Q000378:metabolism*; D019065:Virus Assembly / Q000502:physiology*
chemical_list: D020409:Molecular Motor Proteins; D014764:Viral Proteins; C065038:gp17 protein, Enterobacteria phage T4; D000255:Adenosine Triphosphate