A single protein keeps TB bacteria alive in the lungs - and now scientists can see it

Somewhere inside the lungs of roughly 10 million people who fall ill with tuberculosis each year, a molecular wire is humming. The bacterium Mycobacterium tuberculosis has carved out lipid-rich pockets in damaged lung tissue, and there it feeds - breaking down fats from destroyed cells, channeling the released energy into the machinery that keeps it alive. Cut that wire, and the bacterium starves. But until now, nobody could see the wire clearly enough to know where to cut.

A team at The Hospital for Sick Children (SickKids) in Toronto has changed that. Using high-resolution cryo-electron microscopy, they have produced the first detailed three-dimensional structure of a protein called EtfD, which acts as the critical conduit between fat metabolism and energy production in TB bacteria. Alongside the structure, they developed the first biochemical assay capable of measuring EtfD activity in real time. Together, these tools give researchers what they have lacked for years: a way to screen for drugs that could block TB's survival strategy at its source.

The fat-feeding problem

Tuberculosis kills roughly 1.3 million people annually, making it the leading infectious disease killer worldwide. Part of what makes it so difficult to treat is the bacterium's ability to enter a dormant state inside the lung. Standard treatment requires six months to a year of antibiotics - sometimes longer for drug-resistant strains - and the side effects are punishing enough that many patients struggle to complete the regimen.

The persistence problem traces back to lipids. When M. tuberculosis colonizes the lung, it triggers the formation of granulomas - walled-off clusters of immune cells. Within these structures, the bacterium feeds on fats released by damaged host cells. This lipid diet does more than keep the bacterium alive. It makes it tolerant to antibiotics, slowing its metabolism enough that drugs designed to target rapidly dividing cells lose their punch.

EtfD sits at the heart of this metabolic trick. The protein functions like an electrical wire, ferrying electrons released during fatty acid breakdown into the electron transport chain - the system the bacterium uses to produce adenosine triphosphate (ATP), its primary energy currency. Without EtfD, that energy transfer stalls.

Seeing the wire at atomic resolution

The structural work was carried out at SickKids' Nanoscale Biomedical Imaging Facility. Dr. John Rubinstein, Senior Scientist in the Molecular Medicine program and senior author on the study, led the effort alongside first author Gautier Courbon, a PhD candidate in the Rubinstein Lab.

The cryo-electron microscopy images revealed EtfD's architecture in enough detail to identify where potential drug molecules might bind. That kind of atomic-level map is essential for rational drug design - rather than testing compounds blindly, researchers can now look for molecules shaped to fit specific pockets on the protein's surface.

But a structure alone is not enough. To test whether a candidate drug actually blocks EtfD, you need a way to measure the protein's activity. That is where Courbon's second contribution comes in. The biochemical assay he developed allows researchers to watch EtfD work in real time, seeing precisely when the electron transfer pathway is active and when it is shut down.

From structure to screening

The combination of structure and assay creates what amounts to a starter kit for drug discovery. EtfD had been proposed as a promising drug target before - notably by co-authors Drs. Sabine Ehrt and Dirk Schnappinger at Weill Cornell Medicine - but without a functional assay, there was no practical way to screen inhibitors against it.

Early collaborative work with the SPARC Drug Discovery Facility is already underway. The plan is to test libraries of potential compounds that could block EtfD, looking for molecules that shut down the electron wire without interfering with human metabolism.

The appeal of this target goes beyond novelty. If a drug could cut off lipid-based energy production during the dormant phase, it might dramatically shorten treatment times. Current regimens are long precisely because the bacterium hunkers down in its fat-feeding state and waits out the antibiotic assault. A compound that disrupts that dormancy could make existing antibiotics more effective, potentially reducing treatment from months to weeks.

Mouse models and the road ahead

There are important caveats. This work is structural and biochemical - it establishes a target and provides tools, but no drug candidates have been tested in animal models yet, let alone in humans. The path from a validated target to an approved drug typically takes a decade or more, and most candidates fail along the way.

The rise of drug-resistant TB strains adds urgency but also complexity. Any new drug would need to work against resistant bacteria, not just the standard strains. Whether EtfD inhibitors can clear that bar remains an open question.

Still, the significance of having both a high-resolution structure and a functional assay for a previously intractable target should not be understated. Many promising TB drug targets have stalled precisely because researchers lacked one or both of these tools.

A disease that demands new approaches

TB has plagued humanity for millennia. The bacterium's genome reveals evidence of co-evolution with human populations stretching back tens of thousands of years. Modern antibiotics pushed mortality rates down dramatically in the mid-20th century, but progress has plateaued. The current standard of care - a four-drug cocktail taken for six months or more - has barely changed since the 1980s.

Meanwhile, multidrug-resistant and extensively drug-resistant strains are spreading, particularly in sub-Saharan Africa and Southeast Asia. The World Health Organization estimated in 2024 that roughly 400,000 people developed multidrug-resistant TB in a single year.

Against that backdrop, EtfD represents a genuinely new angle of attack. Rather than targeting the bacterium's growth machinery - the strategy behind most existing antibiotics - it targets the metabolic adaptation that allows the bacterium to survive when growth slows down. That is a fundamentally different approach, and one that could complement rather than compete with existing treatments.

The study was published in The EMBO Journal and funded by the Canadian Institutes of Health Research (CIHR).