Amyloid fibers – long, ropy fibers produced when a functional protein aggregates – are present in the tissues of patients with Alzheimer’s disease, Parkinson’s, type II diabetes, and a number of other diseases. The fibers were long thought to be the causes of these diseases, but amyloid proteins can take many shapes, and research over the past decade has revealed that the amyloid fibers aren’t their most toxic form.
Instead, small clusters of just a few proteins molecules—called oligomers—are now considered more toxic to cells, and likely the form that causes disease. The smaller clumps of protein can cause damage before coalescing into the hallmark fibers, scientists hypothesize.
“This could put amyloid diseases more on the same footing as other diseases.”
Those smaller clumps are typically short-lived, however, and although scientists are increasingly recognizing their contributions to disease, they have been notoriously hard to study. New research from Howard Hughes Medical Institute (HHMI) scientists, published March 9, 2012, in the journal Science, begins to elucidate their role by taking advantage of one amyloid protein that forms fibers more slowly than most. By determining the three-dimensional structure of a segment of that protein, which appears to be a small, toxic oligomer, the researchers are helping researchers understand what causes amyloid diseases and how to design drugs to target the oligomers.
“There are at least 30 diseases lumped into this class of amyloid diseases, and what joins them all together is the involvement of amyloid fibers,” says HHMI investigator David Eisenberg of the University of California, Los Angeles. For almost a century, scientists assumed that these fibers were the cause of amyloid diseases. But when they started isolating the amyloid proteins, they found that the fibers were not the most toxic form. Instead, a handful of labs found, smaller arrangements of the proteins—the oligomers-- could kill cells.
Scientists have begun to link the oligomers of amyloid proteins to disease. In an experiment done in 2010, for example, a team of researchers found that amyloid proteins that could form oligomers but not coalesce into full fibers, were sufficient to cause the clinical signs of Alzheimer’s disease in mice.
When scientists tried to isolate these oligomeric complexes, however, they ran into a roadblock: the amyloid oligomers were incredibly short-lived and unstable. The proteins tended to quickly rearrange into fibers rather than oligomers.
Eisenberg’s lab experienced similar frustrations in working with the amyloid proteins that cause Alzheimer’s disease and one type of diabetes. “You turn your back and they form fibers,” Eisenberg says. Eisenberg’s lab instead focused their attention on a protein that—over many decades—forms amyloid fibers in the lens of the eye, one cause of cataracts. Since the protein, called αβ crystalline (ABC), forms fibers so slowly, they thought it might be easier to coax it to form stable oligomers.
The scientists relied on an antibody—A 11—that’s known to only bind to amyloid oligomers, not the full fibers to study the presence of oligomers. Then, they discovered a segment of ABC that binds with five other copies of itself to form oligomers.
The scientists could also encourage this sticky ABC segment to form fibers by heating the mixture to 50 degrees Celsius and shaking it for a few hours. But at room temperature, the protein formed oligomers. Eisenberg’s team studied the oligomers and found that they were always composed of exactly six copies of the ABC segment.
In their next experiment, the researchers tested whether the oligomers were, in fact, toxic to cells. When they applied the ABC segment in fiber form to cells, the cells stayed alive. But when the ABC oligomers were applied, between 30 and 50 percent of cells died. X-ray crystallography revealed that the six ABC proteins in each oligomer are arranged in a unique cylindrical structure. “If you look through the database of known protein structures, there are no other proteins that look like this,” says Eisenberg. “We named it the cylindrin.”
Eisenberg later showed that the cylindrin is less energetically favorable than the amyloid fibers, explaining why amyloid proteins are usually found in the latter state. But the cylindrin could be key to understanding how amyloid diseases are caused and how they kill cells. The cylindrins could potentially cause channels to form through membranes, for example. More work is needed to discover the role of the six-membered cylinder.
“At this point, it’s still a hypothesis that the toxic forms of these amyloid proteins are cylindrins,” says Eisenberg. “But it’s a testable hypothesis and we can now ask whether other amyloid proteins—those that cause Alzheimer’s of diabetes—also form these structures.”
Knowing the structure, Eisenberg adds, is the first step toward developing drugs to block the cylindrins, if they do turn out to be the disease-causing form of amyloids.
“This could put amyloid diseases more on the same footing as other diseases,” he says.