*Please note: This article was updated on Oct. 15 to include a reference to former President Bill Clinton (D) in the fifth paragraph.
By Daniel Dunaief
David Thanassi wants to give dangerous bacteria in the kidney a haircut.
No, not exactly, but Thanassi, Zhang Family Professor and Chair of the Department of Microbiology and Immunology at the Renaissance School of Medicine at Stony Brook University, has studied how hair-like structures called P pili in the bacteria Escherichia coli are assembled on the bacterial surface.
These pili allow bacteria to hang on to the walls of the kidney, where urine would otherwise flush them out.
Learning about pili at different stages of development could provide a way to keep them from attaching themselves to the kidney and from entering the bloodstream, which could lead to the potentially lethal problem of bacterial sepsis. Indeed, this week, former President Bill Clinton (D) checked into the intensive care unit at the University of California Irvine Medical Center after a urinary tract infection spread to his bloodstream.
“We have been looking at this as a really important aspect of initiating infection from a bacteria’s point of view,” Thanassi said. “How do they build these structures” that lead to infection and illness?
Recently, Thanassi published the structure of these pili in the journal Nature Communication.
The current work builds on previous efforts from Thanassi to determine the structure of these pili in the bladder. He has been exploring how the thousands of proteins that make up the pili get transported and assembled in the correct order. “If we can understand that aspect, we can disrupt their assembly or function,” he said.
Urinary tract infections are a major infectious disease, particularly for women. Indeed, about half of all women will have at least one urinary tract infection, which can be uncomfortable and can require some form of medication.
In some cases, the infections can be recurrent, leading to frequent infections and the repeated need for antibiotics.
The bacteria that cause these infections can become resistant to antibiotics, increasing the importance of finding alternative approaches to these infections, such as interfering with pili.
To be sure, the solution to reducing the bacteria’s ability to colonize the kidney or urinary tract would likely require other steps, as these invaders have additional ways beyond the pili to colonize these organs. Nonetheless, disrupting the way they adhere to the kidney could be a constructive advance that could lead to improved infection prevention and treatment.
One likely strategy could involve using an anti-pilus treatment in combination with other antibiotics, Thanassi explained.
For people who have recurrent infections, anti-pilus therapeutics could offer a solution without resorting to long-term antibiotics.
In his lab, Thanassi is interested in small molecules or chemicals that would disrupt the early stage in pili assembly. “We think of these as protein-protein interactions that are required to build these” pili, he said.
By using a fluorescence reporter, Thanassi and his colleagues can screen libraries of chemicals to determine what might inhibit the process.
As with many biological systems, numerous compounds may seem appropriate for the job, but might not work, as medicine often requires a specific molecule that functions within the context of the dynamic of a living system.
For the helpful bacteria in the gut, pili are not as important as they are for the harmful ones in the kidney, which could mean that an approach that blocked the formation of these structures may not have the same intestinal and stomach side effects as some antibiotics.
To determine the way these pili develop structurally, Thanassi and his lab used molecular and biochemical techniques to stop the assembly of pili at specific stages.
Bacteria assemble these pili during the course of about 30 minutes. An usher proteins serves as the pilus assembly site and pilus secretion channel in the bacterial outer membrane. The usher acts as a nanomachine, putting the pilus proteins into their proper order. A chaperone protein brings the pilus subunits to the usher protein.
In their development, the pili require a protein channel, which is an assembly site.
Thanassi started by working on the usher protein in isolation. The usher proteins function to assemble the thousands of pilus subunits that make up each pilus fiber. The process also involves chaperone proteins, which bind to nascent subunit proteins and help the subunits fold. The chaperone then delivers the subunit proteins to the usher for assembly into the pilus fiber. He used molecular and biochemical methods to express and purify the usher protein.
The assembly process involves interactions between chaperone-subunit complexes and the usher. Over the years, Thanassi has determined how the different proteins work together to build and secrete a pilus.
He was able to force the bacteria to express only one version of the assembly step and then isolate that developmental process.
The majority of the pilus is like a spring or a coil, which can stretch and become longer and straighter to act as a shock absorber, allowing the bacteria to grab on to the kidney cells rather than breaking.
Other researchers are studying how they might make the pili more brittle, preventing that spring-like action from working and compromising its ability to function.
“We’re trying to prevent the pili from assembling in the first place,” Thanassi explained. “Our approach is to try and get molecules that prevent the interaction from occurring.” He is looking at the specific function of one molecule that prevents the usher assembly platform from developing properly, which would wipe out the assembly site.
Thanassi credits former Stony Brook Professor Huilin Li, who is now Chair in the Department of Structural Biology at the Van Andel Institute in Grand Rapids, Michigan, with providing structural insights from his work with the cryo-electron microscoipe. The technology has “revolutionized the work we do,” said Thanassi.
Residents of Smithtown, Thanassi and his wife Kate Kaming, who is Senior Director of Cancer Development at Northwell Health Foundation, have two children. Joseph, 22, attends Northeastern University. Miles, 20, is studying at the Massachusetts Institute of Technology.
Thanassi grew up in South Burlington, Vermont and is an avid skier. He also enjoys mountain biking, walking and music.
Thanassi hopes this latest structural work may one day offer help either with the prevention of infections or with their treatment.
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