A team of scientists from the U.S.
Department of Energy's (DOE) Brookhaven National Laboratory, Harvard University,
and the Indian Institute of Science has made a major step in understanding how
molecules locate the genetic information in DNA that is necessary to carry out
important biological processes. The research, published in the December 1, 2009
edition of Nature Structural + Molecular Biology, confirms that many proteins
responsible for interacting at specific sites on DNA find their targets by sliding
along one of the grooves of the DNA double helix in a spiraling fashion.
Brookhaven biologist Walter Mangel
“Essentially, proteins that search for specific information spin down
the double helix of the DNA, like traveling along the threads of a screw, until
they locate their target,” said co-author Walter Mangel, a Brookhaven
biophysicist.
This research provides experimental proof of a recent theory put forth by the
team and could lead to new ways to alter the behavior of DNA-binding proteins,
which are responsible for replicating and repairing DNA, and for turning genes
on and off.
For decades, scientists have known that proteins searching for genetic sequences
are able to locate them at rates much faster than expected. They found that
rather than moving around the entire three-dimensional space inside a cell,
they moved in one-dimension, along DNA molecules. The Harvard group showed,
in 2006, that the proteins slide back and forth in direct contact with the DNA
as part of the search for specific sequences.
Until now, however, the exact nature of the path these molecules take along
the DNA has not been known. Competing biological models assert that the proteins
either move in a straight line parallel to the DNA axis or trace more complex
helical paths, following a strand or groove of DNA around that axis.
One challenge is that the very fine and quick motions occur at extremely small
space and time scales. This means that the precise motions of a DNA-binding
molecule are difficult to observe directly. So the researchers used indirect
methods to determine the protein's path.
With a special fluorescence microscope, collaborating scientists led by Sunney
Xie at Harvard University observed single protein molecules labeled with a fluorescent
dye binding to and then sliding along the DNA. Although they could not see the
exact path the molecules were sliding on, they could measure how fast the molecules
were going.
Depending on how a protein moves along a DNA axis — either in a linear
or helical pattern —it will encounter different degrees of resistance,
as shown in the earlier paper. If protein motion is linear, its speed will decrease
proportionately as its radius increases. If a protein exhibits helical motion,
it will experience additional friction and its speed will decrease much faster
as its radius increases.
Using a human DNA repair protein as a test for the protein rotation model,
Paul Blainey, now at Stanford University, found the latter case to be true.
When he increased the size of the protein, the rate of motion decreased much
more rapidly than it would have for a simple linear motion.
Relying on the same technique, the group went on to analyze the diffusion rates
of eight different proteins of various sizes. These molecules had highly diverse
functions — such as DNA replication, cleavage, and repair — and
DNA-binding mechanisms. They were also taken from a range of organisms, including
mammals, bacteria, and human viruses.
The researchers observed the same pattern: The speed of each protein decreased
dramatically as its radius increased, as predicted by the theory for helical
sliding.
“The data present strong evidence that proteins seek out targeted DNA
sequences by spinning down the helix rather than linearly sliding along its
axis,” said Biman Bigachi, a co-author from the Indian Institute of Science.
This work validates the new equation for describing and predicting the motion
of protein molecules along strands of DNA with a higher degree of accuracy than
ever before. It enhances the possibilities of future research in understanding
and manipulating the DNA-binding and sliding behavior of proteins.
Said Mangel, “By being able to predict the DNA sliding rate of a protein,
one could alter the size of a protein and thereby alter its sliding rate. For
example, certain viral proteins need to slide along DNA in order to cause infection.
A small protein could be designed to bind to the viral protein to slow down
its sliding rate. This might be a useful means to block a virus infection.”
This research was funded by the National Institute of Allergy and Infectious
Diseases, part of the National Institutes of Health, the National Science Foundation,
and the Department of Science and Technology of India.