During the assembly of many viruses, a powerful molecular motor compacts the genome into a preassembled capsid. Here, we present measurements of viral DNA packaging in bacteriophage φ29 using an improved optical tweezers method that allows DNA translocation to be measured from initiation to completion. This method allowed us to study the previously uncharacterized early stages of packaging and facilitated more accurate measurement of the length of DNA packaged. We measured the motor velocity versus load at near-zero filling and developed a ramped DNA stretching technique that allowed us to measure the velocity versus capsid filling at near-zero load. These measurements reveal that the motor can generate significantly higher velocities and forces than detected previously. Toward the end of packaging, the internal force resisting DNA confinement rises steeply, consistent with the trend predicted by many theoretical models. However, the force rises to a higher magnitude, particularly during the early stages of packaging, than predicted by models that assume coaxial inverse spooling of the DNA. This finding suggests that the DNA is not arranged in that conformation during the early stages of packaging and indicates that internal force is available to drive complete genome ejection in vitro. The maximum force exceeds 100 pN, which is about one-half that predicted to rupture the capsid shell.
Bibliographical noteFunding Information:
Our research was supported by the National Institutes of Health (grants GM-071552 and DE-03606), The Burroughs-Wellcome Fund, The Kinship Foundation, and the Arnold and Mabel Beckman Foundation.