New insights into NLRP3 regulation

November 22, 2022

Prof. Matthias Geyer and lead author Dr. Rebecca Fußhöller (neé Brinkschulte). (picture: David Fußhöller / UKB)


Characterization of the ATP-binding site in human NLRP3

The inflammasome protein NLRP3 is a polymodal sensor of cellular stress. Since it induces cell death through pyroptosis, its activation is strictly regulated. Binding of ATP is considered a prerequisite for NLRP3 activation and inflammasome formation. ImmunoSensation2 members Prof. Matthias Geyer and Prof. Eicke Latz, together with their teams at the University Hospital Bonn, have now further characterized the role of ATP in the regulation of NLRP3. The results were recently published in Communications Biology.


Two consecutive signaling events are required in order to induce the formation of the NLR family pyrin domain containing 3 (NLRP3) inflammasome. In a first step, the transcription of the NLRP3 inflammasome components is upregulated. In a second step, NLRP3 is activated directly on a molecular level. This may be caused by various stimuli, including an increase in intracellular ATP concentration. “The ability of NLRP3 to function as an ATPase is a prerequisite to inflammasome formation,” says Matthias Geyer, Director of the Institute of Structural Biology at the University Hospital Bonn.


Detailed analysis of the ATP-binding pocket

The researchers characterized the amino acid residues involved in ATP-hydrolysis of NLRP3 by site-directed mutagenesis. Based on their findings, a multi-step activation mechanism is proposed. In a first step, NLRP3 is transferred from an autoinhibited into a primed state by post translational modifications. “Only then, the protein is able to take up ATP,” Matthias Geyer explains. “ATP-bound NLRP3 is in the active state, as we call it. From here it can multimerize and form an inflammasome, competent for downstream signaling and cell death.” Alternatively, if the inflammation signal is not strong enough, NLRP3 can hydrolyze the bound ATP and revert to the primed state.

“The active NLRP3 oligomer shows an approximately 14-fold increase in ATP hydrolysis activity compared to the inactive state,” says leading author Rebecca Fußhöller (neé Brinkschulte). On the other hand, a mutagenesis dependent decrease of the protein’s hydrolysis activity could be correlated to a decrease in cellular ASC specking and IL-1b secretion.


ATP-binding affinity responsible for CAPS syndrome?

Cryopyrin-associated periodic syndrome (CAPS) is a rare inherited inflammatory disorder that shows three different phenotypes: familial autoinflammatory cold syndrome, a condition that causes episodes of fever, skin rash, and joint pain upon exposure to cold temperatures. Muckle-Wells syndrome, comprising multiple inflammatory processes such as fever, muscle and joint pain, and hearing loss. And neonatal multisystem inflammatory disease, which is a systemic, inflammatory condition characterized by fever, skin rash, joint pain, and central nervous system symptoms, with an onset during early infancy.


The CAPS disease is elicited by the hyperactive NLRP3 protein, which, as shown in the present study, comes with a decrease in ATP hydrolysis activity. It is characterized by a point-mutation in the predicted Glu-switch region of AAA+-ATPases. The ATP-hydrolysis activity of such mutated NLRP3 protein was found to be significantly reduced, or even halved, in comparison to wild-type protein. “It seems possible that the hyperactive NLRP3 inflammasome activity observed in CAPS patients results from a prolonged ATP residence time inside the binding pocket,” Fußhöller closes. The activated NLRP3 protein would be more likely to multimerize and less likely to hydrolyze ATP and fall back into its primed state. Further investigations are required to fully understand the molecular mechanisms underlying CAPS.



The study was funded by the German Research Foundation (DFG) under Germany’s Excellence Strategy – EXC2151–390873048. The work was further supported by a grant from the Else Kröner-Fresenius-Stiftung to M.G (2014_A203).




Brinkschulte, R., Fußhöller, D.M., Hoss, F. et al. ATP-binding and hydrolysis of human NLRP3. Commun Biol 5, 1176 (2022).



Prof. Dr. Matthias Geyer

Institut für Strukturbiologie

Universitätsklinikum Bonn

Tel. +49-228/287-51400

E-Mail: matthias.geyer(at)