BERKELEY 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
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
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
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