Deciphering the Molecular Mechanism of the Bacteriophage φ29 DNA Packaging Motor
The past decade has seen an explosion in the use of single-molecule approaches to study complex biological processes. One such approach—optical trapping—is particularly well suited for investigating molecular motors, a diverse group of macromolecular complexes that convert chemical energy into mechanical work, thus playing key roles in virtually every aspect of cellular life. Here we describe how to use high-resolution optical tweezers to investigate the mechanism of the bacteriophage φ29 DNA packaging motor, a ring-shaped ATPase responsible for genome packing during viral assembly. This system illustrates how to use single-molecule techniques to uncover novel, often unexpected, principles of motor operation.
Key wordsSingle-molecule manipulation Optical tweezers Viral DNA packaging Molecular motor Ring ATPase
We thank Shelley Grimes, Paul Jardine, and Dwight Anderson for developing the in vitro packaging system and critically reading the manuscript. We thank Gheorghe Chistol, Craig Hetherington, Jeffrey Moffitt, Yann Chemla, Aathavan Karunakaran, Douglas Smith, Sander Tans, Adam Politzer, Ariel Kaplan, Thorsten Hugel, Jens Michaelis, and Steven Smith for their contributions to the development of the single-molecule packaging assay, optical tweezers instrumentation, and data analysis tools. The authors are supported by NIH grants R01GM071552 (to C.B.) and K99GM107365 (to S.L.) and a UC MEXUS-CONACYT doctoral fellowship (to S.T.). C.B. is a Howard Hughes Medical Institute investigator.
- 1.Hetherington CL, Moffitt JR, Jardine PJ et al (2012) Viral DNA packaging motors. In: Goldman YE, Ostap EM (eds) Molecular motors and motility. Elsevier, Oxford, pp 420–446Google Scholar
- 14.Bustamante C, Chemla YR, Moffitt JR (2008) High-resolution dual-trap optical tweezers with differential detection. In: Selvin PR, Ha T (eds) Single-molecule techniques: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp 297–324Google Scholar