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Abstract
ATP-powered viral packaging motors are among the most powerful biomotors known. Motor subunits arranged in a ring repeatedly grip and translocate the DNA to package viral genomes into capsids. Here, we use single DNA manipulation and rapid solution exchange to quantify how nucleotide binding regulates interactions between the bacteriophage T4 motor and DNA substrate. With no nucleotides, there is virtually no gripping and rapid slipping occurs with only minimal friction resisting. In contrast, binding of an ATP analog engages nearly continuous gripping. Occasional slips occur due to dissociation of the analog from a gripping motor subunit, or force-induced rupture of grip, but multiple other analog-bound subunits exert high friction that limits slipping. ADP induces comparably infrequent gripping and variable friction. Independent of nucleotides, slipping arrests when the end of the DNA is about to exit the capsid. This end-clamp mechanism increases the efficiency of packaging by making it essentially irreversible.
Highlights
ATP-powered viral packaging motors are among the most powerful biomotors known
We introduce an assay in which DNA packaging is first initiated with ATP and, after proceeding for a defined time, the actively packaging complex is suddenly moved out of ATP and into solutions containing either no nucleotides, high-concentrations of a non(or slowly) hydrolyzable ATP analog (γ-S-ATP), or the hydrolysis product, ADP
This allows us to characterize the nature of the motor-DNA interactions in defined conditions where motor subunits are switched to a single state: apo, ATP analog-bound, or ADP-bound, respectively
Summary
Motor subunits arranged in a ring repeatedly grip and translocate the DNA to package viral genomes into capsids. A model based on structural data for T4 proposes that positive charged residues in the aligned subunit grip the negatively charged DNA phosphates via electrostatic interactions[20]. One piece of evidence is that when the [ATP] is lowered, slipping occurs more frequently[24,25,26] Another piece of evidence is that pauses in translocation are induced when a small amount of a non- (or slowly) hydrolyzable ATP analog is mixed with ATP. This has been interpreted as gripping of the DNA by an analog-bound subunit[24,25]
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