Stanford Report Online

Stanford Report, October 4, 2000
Bio-X grant recipient relies on chaperones in her lab


"You've got to know when to fold 'em," says the famous country western song.

Sound advice for playing poker -- or making proteins.

It turns out that nearly every molecule of protein produced in your body has to be folded into a specific, three-dimensional shape in order to function properly. Because it translates the linear genetic code into three dimensions, protein folding has been called the second, and more intractable, half of the genetic code.

X-rays of the protein hemoglobin, for example, reveal a complex molecule that resembles a ball of twisted ribbon ­ a unique shape that allows hemoglobin to carry oxygen through the bloodstream. If the molecule is folded incorrectly, oxygen will not be delivered.

Judith Frydman received a Bio-X grant for her research on molecular chaperones.
photo: L.A. Cicero

The human body produces thousands of proteins, each with a distinct function and shape. Some resemble convoluted pretzels, while others are woven into intricate braids.

Biologists used to believe that every protein molecule folded spontaneously as it was being assembled inside the living cell, but recent studies show that many proteins actually need help getting into shape.

Enter molecular chaperones.

Discovered just over a decade ago, chaperones are small proteins that help bigger ones fold and assemble. Some chaperones literally serve as escorts, guiding and twisting newborn proteins into their correct shape.

"No one expected that a cell would require special machinery for protein folding," says molecular biologist Judith Frydman, "but what we are finding is that almost every interesting protein needs a chaperone."

Frydman, an assistant professor of biological sciences, is one of a handful of international researchers whose work has led to new discoveries about molecular chaperones and their vital role in sustaining life.

"Chaperones prevent proteins from folding prematurely or sticking to debris inside the cell," she says. "Without chaperones, some cells get very, very sick."

Frydman's research team is investigating the possible link between chaperones and diseases such as cancer, Alzheimer's and ischemia (insufficient blood supply to the heart and various tissues) that may be caused by the failure of key proteins to fold.

"Understanding how proteins fold in the cell is a central problem in modern biology," she observes, "yet surprisingly little is known about how the process occurs."

The Bio-X Interdisciplinary Initiatives Committee agreed, this week awarding Frydman and her co-researchers a two-year grant to study chaperone-assisted folding.

"It is not known how molecular chaperones perform their 'molecular origami,'" she wrote in her Bio-X proposal.

To solve the problem, Frydman and her research team proposed using a powerful new optical technique called single-molecule spectroscopy, in which lasers, microscopes and ultrasensitive detectors follow one copy of the molecule of interest at a time.

"When this is done," she wrote, "we can actually see how each copy of the protein may march to a different drummer. This is equivalent to watching just one dancer on a dance floor at a time, rather than averaging over all the dancers present which may not be synchronized. New information on how proteins find their native shapes with the help of chaperones will be obtained, which may lead to improved therapies for diseases in which this process has gone awry."

Protein factories

Life would not be possible without proteins. Some fight disease, while others regulate body functions or combine to form various tissues.

"Practically everything done in a cell is done by proteins," notes Frydman.

A protein molecule is essentially a chain of amino acids strung together inside tiny protein factories known as ribosomes that are found in large numbers in all living cells.

A newly formed protein is supposed to fold into its correct shape as soon as it leaves the ribosome and enters the interior of the cell ­ a place fraught with all sorts of dangers.

"The cell interior is a very stressful place where oxidation and heat can harm proteins," says Frydman.

She points out that molecular chaperones were originally known as heat-shock proteins (Hsp), because "their main job seemed to be preventing other proteins from frying in the cell."

By the late 1980s, scientists had discovered that chaperones play a much larger role in the life of a protein. Since then, researchers have identified several distinct families of molecular chaperones.

"One family consists of small chaperones shaped like tiny hands," says Frydman. "Each chaperone grasps the protein's amino acid chain, lets it go, then grasps it again until the protein is properly folded."

Several members of this family, including chaperones Hsp40 and Hsp70, actually work together in a highly organized, protein-folding assembly line.

Another family group includes a larger chaperone known as TRiC, which is shaped like a hollow cylinder. Frydman was the first person to identify the basic composition and function of TRiC, but exactly how it works remains a mystery.

"Somehow," she says, "a protein enters the TRiC cylinder unfolded, then comes out folded less than two minutes later."

Scientists have determined that TRiC chaperones are common in people and other mammals. Estimates are that 10 percent of all mammalian proteins need TRiC in order to fold properly. Another 20 percent bind to the smaller chaperone, Hsp70.

"We're trying to determine when chaperones are needed," says Frydman. "Nobody understands why some proteins require them and others don't."


"What's emerging now," she adds, "is that chaperones appear to be responsible for protein conformation, which is the key to functionality."

"Conformation" refers to the ideal, three-dimensional arrangement of a protein molecule that allows it to assume different shapes inside the cell without breaking apart.

"A protein does a lot of stuff," Frydman comments, "and that requires a structure that is flexible and dynamic."

As an example, she points to actin -­ a rod-shaped protein that helps cells move and muscles contract.

"Actin has to change shapes regularly," says Frydman, "and it can only acquire its unique structure with the assistance of TRiC."

Frydman's research team recently discovered that TRiC also plays an essential role in activating a protein known as VHL, which suppresses a variety of tumors. Without TRiC, VHL will not function, and the consequences can be devastating.

"People who have the VHL defect often develop benign or malignant tumors in the kidney, retina, cerebellum or pancreas," notes Frydman.

"Mutations in the human VHL gene are linked to a hereditary cancer syndrome newly diagnosed in over 20,000 Americans each year," she adds.

"We are currently investigating the role of protein folding in the regulation of VHL activity, and how protein folding is affected in a number of tumor-causing mutations."

Molecular chaperones perform another vital service -­ escorting damaged proteins out of the cell.

"Chaperones decide whether to remove misfolded proteins or to re-fold them," says Frydman. "What's interesting to us is how they make that decision."

Abnormally folded proteins usually have chaperones attached to them, otherwise the proteins tend to clump together in large aggregates similar to the plaques that form on brains of people suffering from various neurodegenerative diseases, including Alzheimer's, Huntington's, Parkinson's and Creutzfeld-Jacob (or mad cow).

Frydman and her colleagues are trying to determine what role chaperones might play in preventing these incurable illnesses.

"All of these may turn out to be diseases of protein conformation," she says. "Clearly, something in the cell has twisted the balance from protein folding to protein aggregation. The fact that people are now looking at molecular chaperones as a possible treatment for Alzheimer's is very exciting for us."