Silent chaperone reveals how TB protein flips immune switch

Tuberculosis remains a global health challenge, and scientists are still working to understand how the bacterium Mycobacterium tuberculosis interacts with the human immune system. One bacterial protein, HtpG of Mycobacterium tuberculosis (HtpG Mtb), helps the pathogen cope with stress by folding newly made or misfolded proteins, acting as an ATP-dependent chaperone alongside other chaperones. Beyond this housekeeping role, HtpG Mtb can also trigger immune responses: it activates Dendritic Cells through a pathway involving the immune receptor TLR4. But until now researchers lacked detailed structural and biophysical information to explain how HtpG Mtb carries out its enzymatic work and how it switches on TLR4. Led by Rita Berisio, the team set out to fill that gap by solving the first crystal structure of HtpG Mtb bound to a non-hydrolysable ATP analogue, and by measuring how the protein interacts with TLR4. Their goal was to link the molecule’s shape and binding behavior to both its catalytic capacity and its ability to engage the immune receptor.

The researchers used crystallography to capture HtpG Mtb in complex with the non-hydrolysable form of ATP, AMPPNP, producing the first detailed structural snapshot of this mycobacterial chaperone. The crystal structure shows that HtpG Mtb forms a dimer that adopts a conformationally silent arrangement; in this state the catalytic domains are positioned so they cannot dimerise in the way required for ATP hydrolysis. In parallel binding studies reported in the same work, HtpG Mtb was shown to bind directly to TLR4 with nanomolar affinity. Importantly, the data indicate that a single HtpG Mtb dimer can engage two TLR4 molecules at once. Taken together, these experimental results link the observed AMPPNP-bound, silent structural state of the HtpG Mtb dimer to a capacity to physically bring TLR4 receptors together, offering a molecular explanation for how the bacterial chaperone activates the immune pathway.

These findings provide a new, concrete picture of how a mycobacterial chaperone both manages protein folding and influences host immunity. Revealing a silent ATP-bound conformation of HtpG Mtb clarifies one aspect of the protein’s catalytic cycle: in this state the chaperone’s catalytic domains are unable to carry out ATP hydrolysis. At the same time, showing that the HtpG Mtb dimer can bind two TLR4 molecules and promote their association suggests a direct physical mechanism for receptor activation that resembles the behavior of LPS, described here as an LPS-like mode. By connecting structure to function, the work offers a platform for future studies that could explore how to modulate HtpG Mtb’s enzymatic activity or its interaction with TLR4. While further work will be needed to translate these structural insights into therapies or vaccines, the study delivers a critical molecular foothold for understanding a dual role of HtpG Mtb in stress response and immune activation.