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The Wonder of Seeing Molecular Machines
The laboratories of David Eisenberg and Todd Yeates are playing a key role in our understanding of complex structures at the molecular level.
When Tony Chan, dean of physical sciences, talks about chemists David Eisenberg and Todd Yeates, he describes them as "molecular ophthalmologists"—scientists who enable us to see significant molecular machines for the first time."You cannot understand the workings of a machine until you know what the parts are and how they fit together," Chan said. "When you can finally see that, then you have a hope of repairing or re-designing a machine that does not work properly. David Eisenberg and Todd Yeates are shining a light on how complex biological machines work by discovering how proteins fit together and revealing their three-dimensional structures."
For Eisenberg, director of the UCLA-Department of Energy Institute of Genomics and Proteomics and a Howard Hughes Medical Institute investigator, "Seeing new structures and finding the unexpected—there's no thrill like it. We are witnessing a revolution in molecular biology and molecular medicine. The day-to-day research is wondrous."
Two aspects of research by Eisenberg and Yeates fit into this revolution: first, they are determining the three-dimensional structures of large, complex proteins using X-ray crystallography—the most powerful tool for discovering the precise arrangement of atoms in a protein. They have determined the structures of more than 100 proteins.
Second, they are learning the rules and patterns for how proteins fold and fit together. The folding patterns that proteins assume are critical to explaining many life processes; for instance, as we age, more of our proteins fold abnormally. Eisenberg and Yeates are learning the underlying principles that govern how molecules assemble into organized structures.
David Eisenberg: Investigating Fundamental Three-Dimensional Structures
Eisenberg and his international team of chemists and molecular biologists discovered a fundamental molecular mechanism that seems to play an important role in Alzheimer's disease, Parkinson's disease, mad cow disease and two dozen other degenerative and fatal diseases. The discovery was featured in the journal Nature in June 2005.
The Eisenberg team discovered the three-dimensional structure—the precise positions of all the atoms—of a miniscule, yet powerful region of a protein that forms dangerous deposits in the brain. The protein is called an amyloid fibril, a rope-like structure created by linked protein molecules that is the common feature of many lethal diseases—and may well hold important clues to treating or preventing them. Determining the molecular structure of a fibril is a feat that eluded researchers for decades.
This particular region of fibril-forming proteins forms sheets that zip together like a surprising "molecular zipper." Eisenberg described this region as "pathologically dry."
"Proteins live in water, but here all the water is squeezed out as the fibril is sealed and zipped up," said Eisenberg. "Our hypothesis is that this dry steric zipper forms in all of these diseases, and is universal in the fibrils. Once this steric zipper has formed, it is very difficult to reverse because it's so tight. We have seen the teeth of the zipper in two related peptides."
Knowing the structure, said Melinda Balbirnie, a UCLA postdoctoral scholar who works in Eisenberg's lab, "may provide a basis for developing drugs to fight these diseases." Balbirnie made the discovery that a small fragment of a protein—a mere one percent of the protein —can behave similarly to the entire protein, and is able to form fibrils. "Like a detective, Melinda traced this fibril-forming property down to a little peptide," Eisenberg said.
Can scientists prevent the steric zipper from forming in the first place, or pry it open once it has formed?
Balbirnie is able to produce fibrils from the small piece of the protein. She is conducting experiments with a wide variety of chemical compounds to see whether any will break up the amyloid fibrils.
"Our Nature paper presents the first atomic-level look at any of these structures," said Rebecca Nelson, a UCLA graduate student in biochemistry and molecular biology.
"We wanted to learn which atomic-level interactions were giving the peptide the property to form the type of fibrils which the body cannot break down," Nelson said. "We thought if we could better understand the structure of the molecules inside the fibrils, we would learn more about why they have the properties they do, how they form, why they might be involved in disease and conceivably how to get rid of them or even prevent their formation."
The researchers discovered that a common feature in these amyloid diseases is that the fibrils can all be characterized by a "cross-beta diffraction pattern" that provides a unique visual signature, said Michael Sawaya, a research scientist with UCLA and the Howard Hughes Medical Institute.
"The fibrils diffract in a way that tells us there are many extended protein chains stacked like a spine or the rungs of a ladder," Sawaya said.
Eisenberg's laboratory has already solved the next piece of the puzzle involving amyloid fibrils. In a November issue of Nature, his UCLA team, which includes graduate student Shilpa Sambashivan and Sawaya, revealed further insights into how amyloids form, and new clues about how to disrupt them. Sambashivan's study also shows that proteins in the amyloid state display common properties.
Todd Yeates: Exploring "Molecular Microcompartments"
Between the publication of Eisenberg's two studies in Nature, Yeates and his team of UCLA biochemists published a major study as well. While Eisenberg's group has been studying proteins that assemble into linear filaments, Yeates' group looks at how some other proteins assemble to form giant, elaborate shells. They discovered the first structural details of a family of mysterious objects called microcompartments that seem to be present in a variety of bacteria—research published in the journal Science in August.
"This is the first look at how microcompartments are built, and what the pieces look like," said Yeates, UCLA professor of chemistry and biochemistry, and a member of the UCLA-DOE Institute of Genomics and Proteomics. "These microcompartments appear to be highly evolved machines. From these first structures, we can see the particular amino acids and atoms."
A key feature that distinguishes the cells of primitive organisms such as bacteria, known as prokaryotes, from the cells in higher organisms like humans is that complex cells—eukaryotic cells—have a much higher level of organization within the cell itself. Eukaryotic cells contain membrane-bound organelles—structures within the cell that perform specific functions. Among these are mitochondria, the tiny power generators in cells. In prokaryotes, cells have been viewed as very primitive, although some contain unusual enclosures known as microcompartments, which appear to serve as primitive organelles inside bacterial cells, carrying out special reactions in their interior.
"Students who take a biology class learn in the first three days that cells of prokaryotes are uniform and without organization, while cells of eukaryotes have a complex organization," Yeates said. "That contrast is becoming less stark; we are learning there is more of a continuum than a sharp divide. These microcompartments, which resemble viruses because they are built from thousands of protein subunits assembled into a shell-like architecture, are an important component of bacteria."
Yeates' Science paper reveals the first structures of the proteins that make up a particular shell called the carboxysome, and the first high-resolution insights into how the carboxysome functions.
"Bacterial microcompartments have remained shrouded in mystery, largely because of an absence of a detailed understanding of their architecture and what the structures look like," said Yeates, who is also a member of the California NanoSystems Institute and UCLA's Molecular Biology Institute.
The structure of the carboxysome shows a repeating pattern of six protein molecules packed closely together. The UCLA biochemists determined the structures from their analysis of small crystals, using X-ray crystallography.
The UCLA biochemists also report 199 related proteins that presumably do similar things in 50 other bacteria, said Yeates, who combines advanced mathematics, chemistry and molecular biology in his research.
Yeates' research team includes lead author Cheryl Kerfeld, director of UCLA's Undergraduate Genomics Research Initiative, Sawaya, graduate student Shiho Tanaka, and UCLA chemistry and biochemistry graduate student Morgan Beeby.
The research could lead to applications in reducing greenhouse gases, Kerfeld said. "The carboxysome is a specialized compartment found in bacteria that 'fix' carbon dioxide," she said. "These organisms can take carbon dioxide and transform it into a form that can be used as an energy source by other organisms.
"A large number of the bacteria in the ocean contains carboxysomes; a major portion of the carbon fixation on Earth is carried out by these naturally-occurring microscopic bioreactors," Kerfeld added. "It's becoming increasingly important to understand how this fundamental process takes place.
"For example, these organisms are known to adapt to a wide range of carbon dioxide concentrations," Kerfeld said. "Studies based on our work could help to reveal how this is accomplished at the molecular level. The carboxysome is truly an elegant design of nature."
Yeates' laboratory will continue to study the structures of microcompartments from other organisms. He also studies the complex relationships among proteins in cells. In addition, he has developed a new strategy for designing novel proteins that self-assemble into a variety of structures or material, including cages, filaments, layers and crystals.
Eisenberg and Yeates are strong advocates of basic research.
"The whole history of science," Eisenberg said, "shows that the solutions to practical problems come from the most unexpected places. We hope our research will show the avenues leading to cures for amyloid diseases, but only time will tell."