Inflammasomes are multiprotein complexes that orchestrate immune responses to pathogenic and sterile insults by regulating the maturation of inflammatory cytokines and pyroptotic cell death. While inflammasome activation is well-characterized at the biochemical level, the mechanisms governing the spatial and temporal assembly of these complexes remain poorly understood. Here, we uncover a critical role for intrinsically disordered regions (IDRs) in activating NLRP1, NLRP3, and NLRP14 inflammasomes. Through structural prediction analyses, we identify IDRs within these receptors that harbor post-translational modification sites essential for inflammasome assembly and function. Notably, disease-associated mutations in NLRP1 and NLRP3 occur within these IDRs, underscoring their functional relevance in inflammatory disorders. Our computational analysis suggests that IDR-mediated phase separation may drive inflammasome condensation at the perinuclear membrane, serving as a sensor for cellular stress, as stress signals may change their conformation, through post-translational modifications, and thus their interaction capacity. Furthermore, inflammasomes lacking IDRs in their NLRPs may rely on interactions with chaperone or adapter proteins containing IDRs for proper assembly. These insights provide a new framework for understanding the regulation of inflammasomes, suggesting that targeting the dynamics of phase transitions could open novel therapeutic avenues for treating inflammatory and autoimmune diseases.
Teresa Nava-Ramírez, C. Cuevas-Velazquez, Alejandra A. Covarrubias et al.· Frontiers in Immunology· 0 citations
The cattle tick Rhipicephalus microplus may rely on an alternative polyamine biosynthesis pathway mediated by arginine decarboxylase activity. This hematophagous ectoparasite is a major constraint to livestock production in tropical and subtropical regions, and its control is increasingly compromised by the rapid emergence of acaricide resistance, highlighting the need for new molecular targets. Previous RNA-seq analyses of ovaries, whole males, and female carcasses identified two sex-enriched transcripts, odc1 and odc2, initially annotated as ornithine decarboxylases (ODCs), enzymes that catalyze the rate-limiting step in polyamine biosynthesis. Polyamines are essential for nucleic acid stabilization, cell growth, stress responses, and gametogenesis. Here, we validated using qPCR that expression of both genes is sexually dimorphic. Phylogenetic analysis across Animalia revealed that R. microplus Odc1 and Odc2 form two divergent tick-specific clades, distinct from canonical ODCs described in mammals and insects. Biochemical characterization of recombinant proteins showed that Odc2 lacked detectable activity under the conditions tested, whereas Odc1 exhibited moderate ornithine decarboxylase activity and robust arginine decarboxylase activity. These results suggests that R. microplus may produce putrescine through an alternative route involving agmatine, thereby bypassing the canonical ornithine decarboxylation pathway. To our knowledge, this represents the first biochemical evidence consistent with arginine decarboxylase activity in an animal enzyme. Together, these findings support the existence of a previously unrecognized branch of polyamine metabolism in ticks and identify Odc1 as a promising candidate target for the development of selective acaricides.
R. Cossío-Bayúgar, E. Miranda-Miranda, Michael F. Dunn et al.· Scientific Reports· 1 citation
The CldA enzyme is an unprecedented functional intermediate exhibiting the dual hydrolytic specificity of starch hydrolases and the intramolecular transglycosylation capacity of cyclomaltodextrin glucanotransferases (CGTases) from subfamily 2 of family 13 of glycoside hydrolases (GH13_2). Here, the crystallographic structure of CldA was determined at 1.66 Å resolution. Structural and kinetic studies revealed that the thermophilic CldA has a three-domain ABC architecture similar to that of starch hydrolases from GH13_1, and it contains three calcium-dependent folding centers (Ca+2 1-3) essential for thermostability. However, it simultaneously features nine expanded subsites (-7 to +2) defining the active site cleft of the canonical five-domain ABCDECBM20 CGTases from GH13_2. Structural comparisons revealed three evolutionary adaptations in CldA: (a) the absence of the substrate-guiding DECBM20 domains; (b) an unusual hydrophobic pair, Trp204/Met281, whose hydrophobicity was critical for stabilizing the cyclodextrin (CD) ring at the acceptor subsite +2; and (c) an unexpected hydrogen bond of 2.60 Å between Ser200 at subsite -6 and the key central aromatic residue, Phe216, involved in starch circularization for CD formation. Characterization of two mutants, CldAM281F and CldAS200G, and a five-domain chimera, CldA-DECBM20, provided insights into the boundary between starch hydrolases and CGTases. CldA provides the first experimental structural evidence for a native GH13_2 enzyme where hydrolytic and cyclization activities coexist at a presculpted CGTase active site. Overall, structural and functional analysis of CldA showed that it diverges from canonical GH13_2 CGTases by lacking C-terminal DE domains, shifting its intramolecular transglycosylation specificity toward hydrolysis through an intriguing starch concentration-dependent product-length mechanism.
Beatriz Velazquez-Cruz, Montserrat Romero-Jiménez, Yasel Guerra et al.· The FEBS Journal· 0 citations
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