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Molecular Motor Packs DNA Into Viruses At Greater Than Champagne Pressures, Researchers Report

Date:
October 19, 2001
Source:
University Of California - Berkeley
Summary:
The DNA inside some viruses is packed so tightly that the internal pressure reaches ten times that in a champagne bottle, according to new measurements by biophysicists at the University of California, Berkeley, and the University of Minnesota.
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The DNA inside some viruses is packed so tightly that the internal pressure reaches ten times that in a champagne bottle, according to new measurements by biophysicists at the University of California, Berkeley, and the University of Minnesota.

The researchers suspect that this high pressure helps the virus spurt its DNA into a cell once it has latched onto the surface. Once the DNA gets inside, it begins retooling the cell to manufacture new viruses. The process eventually kills the cell, but not before generating thousands more viruses to spread the infection.

Such tight packing is achieved by one of the most powerful molecular motors ever observed, stronger than the motors that move our muscles or the nanoscale molecular motors that duplicate DNA or transcribe it into RNA. The motor the researchers studied is part of the bacteriophage ø29 (phi-29), a virus that is the scourge of the common soil bacterium Bacillus subtilis.

"Pound for pound, this is stronger than any known molecular motor, and can pack DNA to a pressure of about 60 atmospheres," said biophysicist Carlos Bustamante, professor of physics and of molecular and cell biology in the College of Letters & Science at UC Berkeley. A bottle of champagne typically is under pressure of five to six atmospheres, the equivalent of nearly one hundred pounds per square inch.

"Many human viruses, such as the herpes viruses that cause herpes simplex, chicken pox and shingles, are thought to pack their DNA in the same way, so understanding how this 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," said Bustamante, who also is an investigator in the Howard Hughes Medical Institute at UC Berkeley and a researcher in the Physical Biosciences Division of Lawrence Berkeley National Laboratory.

Adenoviruses, popular today with gene therapists as vehicles for ferrying genes into cells, also are suspected to pack their genes this way.

Bustamante and his colleagues report their findings in the Oct. 18 issue of Nature. His coauthors are Douglas E. Smith and Sander J. Tans, both former postdoctoral fellows at UC Berkeley; Shelley Grimes and Dwight L. Anderson of the University of Minnesota Departments of Microbiology and Dentistry; and UC Berkeley research associate Steven B. Smith.

Douglas Smith now is an assistant professor of physics at UC San Diego. Tans is at the Institute for Atomic and Molecular Physics in Amsterdam.

Bacteriophages are viruses that attack and kill bacteria, and are composed of a hard shell or capsid containing tightly coiled DNA. They typically glom onto the bacterial surface and inject their DNA into the cell interior. Once there, the DNA takes over the replication machinery to make thousands of copies of the virus, filling the cell until it bursts.

The first piece of the virus made is the empty capsid with a protein complex, called the portal motor, at the mouth. This motor grabs hold of the viral genome, a double strand of DNA, and pushes it into the capsid to complete assembly of the virus.

Bustamante suspected that the motor generates a strong force since it compacts DNA nearly 6,000 times its normal volume. To achieve this, the motor has to overcome DNA's resistance to bending, the electrostatic forces of repulsion encountered when pushing charged atoms close together, and the forces of entropy that make anything resist being constrained in a tight space.

For nearly four years, Bustamante and his colleagues at the University of Minnesota tried pulling on the DNA as it is being stuffed into the capsid in an attempt to measure the force generated by the packing motor. They used a technique Bustamante and Steven Smith helped develop nine years ago to manipulate single molecules.

The technique involves attaching a microscopic bead to the free end of the DNA and attaching a second bead to the capsid. While immobilizing the capsid bead with a pipette, they then capture the bead on the free end in an optical tweezer, which is essentially a laser beam that allows them to pull on the bead at the same time as they measure the resistance.

At first they pulled too hard, stalling the motor and probably stripping its gears. Eventually, the researchers were able to measure the force generated by the motor throughout the entire packing process, allowing them to calculate the total work involved, the total internal pressure and the energy released when the capsid is uncorked.

At maximum, the motor pulled with about 57 to 60 picoNewtons of force. Scaled up to human dimensions, this would be enough to lift six aircraft carriers, Bustamante said. A Newton is a force roughly equivalent to the weight of an apple on Earth, and a picoNewton is a millionth of a millionth of a Newton.

For comparison, the molecular motor RNA polymerase, which creates strands of RNA, exerts a maximum force of 15 to 20 picoNewtons. A similar motor, DNA polymerase, which creates strands of DNA, stalls at 35 picoNewtons, while myosin, the motor that contracts muscle fibers, individually can generate only five picoNewtons of force.

"Fifty-seven picoNewtons is an enormous force," Bustamante said. "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."

The motor is also very efficient. Given sufficient energy in the form of ATP (adenosine triphosphate), the motor chugs along until all the DNA is packed into the viral capsid, with only occasional pauses.

Among the other questions Bustamante and his colleagues hope to answer is whether the bacteriophage's portal motor is a rotating motor, like the motor that powers the tail or flagellum of some bacteria.

"The motor, consisting of a 10-nanometer diameter ring of RNA molecules sandwiched between two protein rings, is very intriguing and different from other motors that have been studied," said Douglas Smith. "We suspect that rotation of the rings may pull the double helical DNA through the portal similar to the way a rotating nut can pull on a bolt."

They also hope to determine once and for all whether the virus injects its DNA into cells by mere passive diffusion, or whether, as they suspect, it uses the packing energy to actively inject its DNA.

The work was supported by the National Institutes of Health, the Department of Energy and the National Science Foundation.

Bustamante is a member of UC Berkeley's Health Sciences Initiative, a broad effort bringing together researchers from many disciplines to work on health problems of the 21st century.


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Materials provided by University Of California - Berkeley. Note: Content may be edited for style and length.


Cite This Page:

University Of California - Berkeley. "Molecular Motor Packs DNA Into Viruses At Greater Than Champagne Pressures, Researchers Report." ScienceDaily. ScienceDaily, 19 October 2001. <www.sciencedaily.com/releases/2001/10/011019074640.htm>.
University Of California - Berkeley. (2001, October 19). Molecular Motor Packs DNA Into Viruses At Greater Than Champagne Pressures, Researchers Report. ScienceDaily. Retrieved March 27, 2024 from www.sciencedaily.com/releases/2001/10/011019074640.htm
University Of California - Berkeley. "Molecular Motor Packs DNA Into Viruses At Greater Than Champagne Pressures, Researchers Report." ScienceDaily. www.sciencedaily.com/releases/2001/10/011019074640.htm (accessed March 27, 2024).

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