June 16, 2014
By C.J. Janovy
|Mark Fisher (left) with Subhashchandra Naik|
A University of Kansas School of Medicine researcher is leading a multi-institutional team of scientists in designing new ways to identify potential new drugs to block the anthrax toxin molecules from causing cell death.
Although human and animal vaccines for anthrax have existed for more than a hundred years, concern about anthrax has been renewed in the post-9/11 era of heightened public awareness surrounding biological and chemical weapons. In nature, anthrax endospores can remain dormant in soil for decades, infecting plant-eating animals through ingestion, inhalation, or injured skin. In 2011, anthrax-laced letters were sent to several national media outlets and government offices in a bioterrorist act that claimed five lives, including two postal workers and a hospital employee.
Mark Fisher, professor of biochemistry and molecular biology, is working with postdoctoral fellows Narahari Akkaledevi and Srayanta Mukherjee at KU Medical Center and Wonpil Im, associate professor in bioinformatics at KU's Lawrence campus. Also collaborating on the project are Ed Gogol, associate professor in biophysics at University of Missouri-Kansas City; R. John Collier, professor of microbiology at Harvard; Brad Pentelute, assistant professor in the chemistry department at MIT; and Steve Ludtke, professor of biochemistry and molecular biophysics at Baylor. The team has built the first structural images of the anthrax toxin component. Called protective antigen pore translocon, it is a unique syringe-shaped protein that injects the other anthrax toxins into cells. See Movie 1, structure transition constructed by Srayanta Mukherjee and Yifei Qi:
As anthrax bacteria set up shop inside the host cell, they produce and release three proteins that combine to form the anthrax toxin complex. In its initial stages, the protective antigen is in a "prepore" state that behaves like a Trojan horse, carrying two lethal toxins that bind to proteins on the surface of the cell. As soon as this prepore is attached, it's engulfed by the cell. Once it's bound to the membrane of the endosome, different acidic levels cause the prepore to change its shape to resemble a "protein needle," punctures the endosome membrane and injects two lethal toxins into the cell interior. These toxins proceed to disrupt cellular function.
To observe the prepore-to-pore transformation process, Fisher and his KU Medical Center colleagues, led by graduate student Subhash Naik, came up with a way to zero in on the movements of the anthrax toxin complex at the molecular level. Using a special instrument that measures changes in light wave interference patterns that can detect molecular changes, Fisher and his team are able to follow the entire assembly and transition process in real time.
"It turns out that the anthrax toxin complex seems to exert its most lethal effects on the heart and liver cells. In addition, anthrax toxins kill protective immune-system cells," he says. "It's an insidious little protein-based lethal injection system." See Movie 2, constructed by Janet Iwasa, Brad Pentelute and R. John Collier, Harvard:
The ability to track and visualize these structure changes in the toxin complex, Fisher says, will allow researchers to develop and test "rational" drugs that can target these toxin proteins directly as they change, reassemble in different shapes, and bind to cells. "It is more than likely that our ability to watch assembly and transitions of other toxins at the molecular will definitely be applicable to other bacterial and viral invasion systems," Fisher explains.
"With vaccine development, the host must mount a massive immune response to develop specific antibodies against the protein toxins, which in turn prevents these toxins from functioning properly," Fisher says. "However, vaccine development can be slow because producing the right formulation of inert toxin proteins is tedious." Another line of defense is antibiotics, although that approach is inefficient, Fisher says. "Antibiotics directly target the bacteria, but the toxins that are released from the bacteria during the infection phase are still present and continue to kill cells directly," Fisher says. "Our approach of directly constructing novel antimicrobial agents to inhibit or prevent toxin assembly and transitions will complement vaccine development and antibiotic treatment efforts."
Ultimately, Fisher says that this technology platform will provide drug validation opportunities to interrupt or block the transition and assembly of these protein toxins. This advance will give physicians and health workers a new and rapid frontline defense against anthrax toxicity.
Fisher's research was published and highlighted last year in the journal Biochemistry and as the cover article in Protein Science. He is also the faculty advisor for Blue Valley High School students and their science teachers, who are working on a project where they print 3D structure models of the anthrax toxin proteins. "This hands-on model approach is a great teaching tool for students and the general public," Fisher says, noting that the students presented their work at this year's American Society of Biochemistry and Molecular Biology annual meeting in Washington D.C.