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Mighty Biomolecular Motors
    In a development with ramifications for gene therapy and infection-fighting drugs, a research collaboration led by Carlos Bustamante of Berkeley Lab's Physical Biosciences Division (PBD) has discovered the mechanism by which at least some viruses inject their DNA into the cells of other organisms. The mechanism involves one of the most powerful biomolecular motors ever observed.

This motor pulls with about 57 to 60 piconewtons of force which, scaled up to human dimensions, would be enough to lift six aircraft carriers," says Bustamante, a biophysicist who, in addition to his affiliation with Berkeley Lab, also holds appointments with UC Berkeley and the Howard Hughes Medical Institute.

Biomolecular motors are proteins that undergo shape changes in order to generate force or torque. Acting like tiny engines, biomolecular motors come in a wide assortment of varieties and perform a broad range of tasks, many involving movement and transportation. One such task is the packing of coiled lengths of DNA into the protective external shell or "capsid" that is prominent on a number of viruses including those that cause herpes, chicken pox and shingles. The biomolecular motor that Bustamante and his colleagues observed is the portal motor for the bacteriophage ø29 (phi-29), a virus that infects and destroys soil bacteria, and is considered an excellent model system for studying viral assembly.

"This motor pulls with about 57 to 60 piconewtons of force which, scaled up to human dimensions, would be enough to lift six aircraft carriers."

"The portal motor for bacteriophage ø29 compresses the DNA into a space that is 6,000 times smaller than its normal volume," says Bustamante. "This generates an internal pressure of about 60 atmospheres, which is about ten times that in a champagne bottle."

Bustamante and his collaborators propose that just as the internal pressure in a
champagne bottle will pop a champagne cork, so too does the even greater internal pressure inside the bacteriophage's capsid forcibly pop the viral DNA into an attacked cell. Viruses cannot "live" or reproduce without getting inside a living cell, whether it's a plant, animal, or a bacterium. In the case of ø29, the bacteriophage attaches itself to and injects its DNA into a soil bacterium, which, unlike the virus, can reproduce on its own. The viral DNA takes over the bacterium's reproductive programming and instructs it to reproduce copies of the virus instead. So many copies of ø29 are replicated that the bacterium ultimately bursts open, unleashing a mass of new ø29 viruses ready to infect other bacteria.

Biophysicist Carlos Bustamante with the optical tweezers setup used to measure the strength of bacteriophage ø29's portal motor.  

"Understanding how this DNA packing process works could help us design better drugs to interfere with the packing part of the infection cycle of the virus and perhaps halt infection," Bustamante says. "It might also be used in gene therapy as a means of transporting new genetic material into cells."

To measure the strength of bacteriophage ø29's portal motor, Bustamante and his collaborators used force-measuring optical tweezers. Working with capsids that were only partially packed with DNA before the packing process was stalled, they tethered the unpacked end of the DNA and the capsid into which it was being packed between a pair of micron-sized polystyrene beads. While the capsid-attached bead was held in place by a pipette, the DNA-attached bead was captured by the optical tweezers-a laser beam that can be used to grasp and move the beads.

  The ø29 motor (yellow) compresses coiled lengths of DNA into the viral capsid to 6,000 times its normal volume, creating pressure 10 times as powerful as that inside a champagne bottle.

In the presence of adenosine triphosphate (ATP), the fuel that powers many biomolecular motors, Bustamante and his collaborators were able to observe viral DNA-packing activity in real time and measure the force being applied by bacteriophage ø29's biomolecular motor. This enabled them to calculate the total amount of work involved, the total internal pressure on the DNA, and the amount of potential energy available for ejecting the DNA out of the capsid and into a bacterium during infection.

"The 57 to 60 piconewtons we calculated as the maximum pull exerted by this motor is an enormous force," Bustamante says. "The question is then, what happens to all the work done on the DNA during packing? We claim the energy gets stored up inside the head of the bacteriophage and becomes available to initiate rapid injection of the DNA during the next infection phase."

Collaborating with Bustamante on this research were Doug Smith, now with UC San Diego, and Sander Tans, now at the Institute for Atomic and Molecular Physics in Amsterdam, along with UC Berkeley's Steven Smith, and Shelley Grimes and Dwight Anderson of the University of Minnesota.

"I would like to emphasize the close collaboration between my laboratory and that of Dwight Anderson that made possible this work," says Bustamante. "It was only because of the excellent complementary expertise of the two laboratories that this phase of the work was successfully completed."

The work was funded by DOE Office of Science, the National Institutes of Health, and the National Science Foundation.

-- Lynn Yarris

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