Uncovering the ATP-Dependent Activation Mechanism of NLRP3 through Integrated Computational Approaches

As a central component of NLRP3 inflammasome, the pattern recognition receptor NLRP3 (nucleotide-binding oligomerization domain (NACHT)-, leucine-rich repeat (LRR)-, and pyrin domain (PYD)-containing protein 3) plays a critical role in immune responses. Its abnormal activation is closely associated with various inflammatory diseases, positioning it as a valuable therapeutic target. During the transition from inactive to active states, NLRP3 undergoes significant conformational changes, including spatial rearrangement of the NACHT domain and an approximately 85.4° rotation of the WHD-HD2-LRR module caused by the rearrangement. These substantial conformational transitions, whose molecular mechanisms remain elusive, present significant challenges for the development of NLRP3 inhibitors. Here, computational approaches including nudged elastic band (NEB) simulations, molecular dynamics (MD) simulations, and Markov state model (MSM) are integrated to investigate the activation process of NLRP3. Our analyses reveal how ATP binding initiates a cascade of conformational changes that propagate throughout the NLRP3 architecture, driving its conformational transition, which establishes a foundation for understanding the NLRP3 activation mechanism. Meanwhile, five representative metastable conformational states, including three key intermediates, are identified, which provide a structural basis for developing novel allosteric inhibitors with improved specificity and efficacy for NLRP3-mediated inflammatory diseases.