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“Smuggler” Protein Finding Could Be Used to Disrupt Bacterial Cell Walls

Stopping bacteria from maintaining their cell walls could lead to new ways to treat infections


New research seeks to identify the unknown protein responsible for ferrying the raw materials bacteria use to build their cell walls.
Credit: NIAID via Flickr

This is the story of a treacherous border crossing, valuable cargo and tracking down a secretive smuggler whose identity has been shrouded in mystery—and controversy—for decades.
 
The harsh border is the hard-to-penetrate membrane surrounding bacteria, which lets few things pass. But the smuggler, a bacterial protein, ferries critical raw material through this barrier to help build the organism’s protective sheath, its cell wall. Stopping the protein should weaken that cell wall—bad for the bacteria but good for patients hosting bacterial infections, because this could lead to new antibiotics.
 
To stop it, however, scientists have to identify the smuggler protein—until recently they have been stymied in their efforts to figure out which of many molecules it is. It has a generic name, a flippase, but that is all. New research, announced in the July 11 Science, attempts to rip the mask off the elusive molecule and dubs it MurJ. The work, led by scientists at The Ohio State University, is the first time microbiologists have caught the protein in the act in living bacteria. But their discovery has also fueled a fierce debate, because other scientists have come up with evidence fingering a different culprit.
 
The bacterial cell membrane serves as a flexible barrier that keeps the cell’s contents in place. Beyond it, the cell wall is a stiff mesh that keeps the membrane from exploding due to enormous internal pressure. The wall is like a chicken wire fence that strengthens and supports the membrane, says Kevin Young, a microbiologist at the University of Arkansas for Medical Sciences. “It’s just as though you took a balloon and encased it in some meshwork and tried to pump air into it. The balloon would not explode.” Young, not a part of the research, described the results as important and talked about the still-hot identity debate in an accompanying article in the same issue of Science.
 
The flippase is important because it helps build this wall from a sugary molecule called peptidoglycan that is manufactured inside the cell. To get to the wall peptidoglycans must pass through the inner membrane, which is made of oily molecules called lipids that repel them.
 
So the flippase sneaks peptidoglycans out without giving the membrane a chance to repel it. How? Imagine putting your hands together so that just your fingertips are touching, says Natividad Ruiz, a microbiologist at Ohio State and a member of the new study’s research team. Now switch, so instead of touching at your fingertips your hands connect at the heels of the palms. By sticking an object in between your hands and switching the opening, you could move the object from palm to fingertip without ever exposing it to anything—like a repulsive membrane—to your left or your right.
 
Ruiz’s team found MurJ doing something very much like that in Escherichia coli bacteria. They inhibited the protein and found that when it was crippled, peptidoglycans do not get across the membrane. This result, along with a computer model of MurJ’s structure, suggests to Ruiz that MurJ is the flippase. “If you look at all of the previous evidence...you have to conclude that MurJ is the protein itself that does this job,” she says.
 
Eefjan Breukink says he does not have to conclude that at all. Breukink, a biochemist at Utrecht University in the Netherlands, favors another protein called FtsW as the flippase. In research published in 2011 in The EMBO Journal the scientist conducted a test tube experiment using extracted bits of bacterial membranes to show that FtsW—not MurJ—had an effect on how much peptidoglycan crossed the border. Breukink says that there are many other potential reasons why Ruiz’s result would indicate MurJ is the flippase. “That’s the difficulty [in living cells]. You always have alternative explanations. We have to purify the protein and show it has that activity [outside the cell].”
 
To Ruiz, however, that is precisely the problem with Breukink’s experiment: It is so artificially removed from the realities of a living cell that its results must be taken with a grain of salt. “They never went back to the cell to test it,” she says. “Ultimately what we want to know is how this works in the cell. So let’s start from the cell, let’s learn within the biological system as much as we can.”
 
The argument is as much about differing scientific philosophies as it is about a pair of proteins, pitting the inherent complexity of living cells against the simple—perhaps oversimplified—world of purified experiments with fewer molecular players. Ultimately, settling the debate will require more than just another experiment in MurJ’s or FtsW’s favor, it will take a result that manages to explain each team’s findings simultaneously while pointing to a single culprit. “We want these two things to come together somehow,” Young says. “What you’d like is for both approaches to give you the same result.”

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