Repulsive DNA-DNA Interactions Accelerate Viral DNA Packaging in Phage Phi29

We use optical tweezers to study the effect of attractive versus repulsive DNA-DNA interactions on motor-driven viral packaging. Screening of repulsive interactions accelerates packaging, but induction of attractive interactions by ${\text{spermidine}}^{3+}$ causes heterogeneous dynamics. Acceleration is observed in a fraction of complexes, but most exhibit slowing and stalling, suggesting that attractive interactions promote nonequilibrium DNA conformations that impede the motor. Thus, repulsive interactions facilitate packaging despite increasing the energy of the theoretical optimum spooled DNA conformation.

Figure 1

(a) The prohead-motor complex is attached to one optically trapped microsphere (left) and the end of the DNA is attached to a second trapped microsphere (right). (b) Typical measurements with 5 pN external load in the standard condition (black), low spermidine (blue, slightly faster), and high spermidine (red, highly heterogeneous).

Figure 2

(a) Average packaging rate and (b) motor velocity (packaging rate not including pauses and slips) versus prohead filling under 5 pN applied force in the standard condition (black, 3rd highest initial rate), low spermidine (blue, 2nd highest initial rate), high spermidine (red, lowest initial rate), and for the subset of complexes in high spermidine that package to $>80\%$ filling (green, highest initial rate).

Figure 3

(a) Average motor velocity versus applied load for complexes that reached $>30\mathrm{pN}$ with $<20\%$ filling in the standard condition (black, $\mathrm{N}=113$ events, lowest rates), low spermidine (blue, $\mathrm{N}=61$), and fast (nonstalling) complexes in high spermidine (green, $\mathrm{N}=101$, highest rates). (b) Standard deviation in DNA length packaged by different complexes for the standard condition (black), low spermidine (blue, nearly equal to standard), and high spermidine (red, highest values) under 5 pN load.

Figure 4

(a) Example where a complex was packaging under 5 pN applied load (shortening of the DNA extension; section no. 1) and was relaxed to bring the applied force to near zero (section no. 2). The DNA was then stretched again to 5 pN (no. 3) and found to be partly condensed (indicated by 4) (dashed line shows the expected trend for packaging in the absence of condensation). Despite condensation, packaging continued (no. 5). (b) Example in which motor was packaging under 5 pN load (no. 1) and then stalled (no. 2). The DNA was then stretched to 25 pN to extend any condensed segments (no. 3), but no recovery in packaging was observed (no. 4). Inset shows standard deviation in lengths measured at 15 pN in the standard condition (black, lower values) and high spermidine (red, higher values).