Coiled strand of the protein chromatin is revealed in this image from the laboratory of Carlos Bustamante of the University of Oregon. He and other researchers are pioneering a new generation of "nanoscopy" techniques that not only take pictures of tiny organic molecular structures, but can manipulate them as well.
Ever since Antonie van Leeuwenhoek peered through his crude microscope and confirmed that life is indeed packaged in very small containers, biologists have recruited physicists to focus stronger lenses, train more powerful rays and run ever tinier probes on cells and the molecules that make them live--and die. Yet many mysteries of life have remained unscrutinized by human eyes. Into what shapes do vital proteins fold themselves? What molecular tricks do viruses such as HIV use to avoid destruction while doing their damage? How is it exactly that cells translate some sections of DNA into proteins yet completely ignore other sections?
Biologists have arrived at educated guesses and rough answers from indirect chemical evidence. Direct pictures would be extremely useful, but seeing the intimate details of biology--a nanoscopic folded protein, a virus's coat, or a DNA helix as it is unwound and transcribed into RNA--requires an instrument that can focus tightly on molecules in their natural, wet environment. Diffraction blurs the vision of optical microscopes at scales below a micron (a thousandth of a millimeter) or so. Electron microscopes capture clear images only of frozen, metal-coated samples. Scanning tunneling microscopes can reveal individual atoms, but only if they conduct electricity well.
In mid-March, at the American Physical Society's annual meeting in Kansas City, Mo., researchers announced success with two newer kinds of instruments. The first, called an atomic force microscope (AFM), is already proving a powerful tool for molecular biologists. The instrument uses a carefully engineered cantilever with a tip just nanometers (millionths of a millimeter) across to scan a surface in the way the needle of a phonograph does. Laser light bouncing off the tip records its movements and traces whatever surfaces it encounters, wet or dry.
Of course, dragging even a tiny a needle across a DNA molecule can damage the molecule or distort its shape. So instead, as Neil H. Thomson of the University of California at Santa Barbara reported, he and his colleagues vibrated the tip, tapping it ever so gently at nanometer steps across the field of view.
And what a view. The researchers saw a remarkable sight: a strand of DNA transcribed by a little glob of the enzyme RNA polymerase into a genetic RNA message, ready to be translated into a protein. The movies they made show that the enzyme does indeed run along the unwound DNA helix like a boxcar on a track, just as biologists have long imagined (but did not until recently know for certain). The technique should allow experimenters to see what really happens when they insert codes into the DNA that should derail the process. A clearer picture of the events should help in designing drugs that switch off genes that cause disease and switch on those that fight it.
Carlos Bustamante of the University of Oregon put the AFM to a different use. Instead of scanning across proteins to see how they behave, he took a more direct approach. He attached one end of the protein titin, thought to be a key player in muscle elasticity, to the tip of his cantilever and attached the other end to a plate. Then he pulled. And pushed. And pulled again, over and over, measuring all the while the force that the protein exerted on the cantilever.
What he found surprised him. The titin molecules he studied did not behave like a rubber band, stretching evenly until they could stretch no more, then shrinking and exerting as much pull as when stretched. Instead they behaved like a rubber band wound up into knots, expanding with little resistance at first but then stretching in jerks and fits. They contracted the opposite way: sharply at first, then with only a little pull. "When we overlay this elasticity profile on top of that for muscle cells, they look identical," Bustamante beams. "It was known that titin played a role in muscle tone, but now we conclude that it in fact controls muscle tone." That finding, if true, could suggest new kinds of drugs for dystonia and other muscular diseases. But the experiments still need to be replicated and verified by others.
Meanwhile, using a technique very similar to the Oregon team's, a group lead by Gil Lee at the Naval Research Lab has measured the force needed to tear apart two complementary strands of DNA. It does not require much oomph--about 500 piconewtons to rip apart 20 base pairs. (An analogy clarifies how little force this is: the weight of your forearm on a desk overpowers 500 piconewtons by about as much as the weight of Hoover Dam outpowers your forearm.)
Yet physicists are working on an even more sensitive device. Dan Rugar and his colleagues at the IBM Almaden Research Center announced on March 17 that they have designed a way to measure magnetic forces as small an attonewton--500,000,000 times smaller than the tug of 20 DNA base pairs. They are now incorporating the new sensor into a magnetic resonance force microscope (MRFM), which combines the strategies of atomic force microscopy and of magnetic resonance imaging, now widely used in medicine.
Although this newer type of microscope has reached only micron-level resolutions, the technology is progressing quickly. "The ultimate goal is to be able to create three-dimensional images of materials in slices as small as one atom wide," says Zhenyong Zhang of Los Alamos National Laboratory, who also has an MRFM to play with. Because this kind of instrument can, like the MRI scanners in hospitals, see inside materials, it promises to one day reveal the sticky crevices in proteins that seem to be so critical in determining their function. Leeuwenhoek would be impressed.