Fig. 2: Motors containing inactive subunits exhibit impaired, but not abolished, activity.

a Schematic of in vitro bulk packaging assay. After allowing the packaging to proceed, the unpackaged DNA was digested by DNase I and the packaged DNA was retrieved from the capsids by digesting the capsid and DNase I with proteinase K. b Agarose gel electrophoresis of DNA packaged by motors constituted with increasing numbers of inactive subunits. The WT gp17 and the inactive Q143A mutant gp17 (inactive subunit) were pre-mixed at different ratios and added to the reaction mixtures containing capsids, ATP, and linearized plasmid DNA. The uncut gel image is shown in Supplementary Fig. 14. c Schematic of single-molecule fluorescence assay. The assay is similar to the one in Fig. 1a but performed with increasing percentage of inactive subunits (black sphere). d No. of Cy5 spots with increasing percentage of inactive subunits. In the inset are representative CCD images of the Cy5 channel, showing the number of Cy5 spots (each containing 1 to n # of packaged DNA) indicate the packaging efficiency with increasing percentage of inactive subunits. e DNA packaging activity quantified from the agarose gel in b (cyan squares). Also shown are predicted activities where the minimum number of active subunits required for packaging is 1 (no coordination; black) to 5 (strict coordination; gray)). f Representative single-molecule trajectories of Cy5 photobleaching steps (red), indicating the number of DNA molecules packaged inside the capsid, and Cy3 photobleaching steps (green), indicating the number of inactive subunits in the motor. Inset: Schematic of the assay, similar to the one shown in c, with inactive subunits (black sphere) labeled with Cy3. g Distribution of the counts of packaged DNA molecules from motors reporting 0–3 Cy3 photobleaching events (indicative of the count of inactive subunits). The average number of packaged DNA is indicated by f. The number of trajectories used is indicated by n = #.