D3.70 - Structural and immunological insights into arginine kinases from house dust mites and related arthropods

Arginine kinases (AKs) are highly conserved enzymes and well-established pan-allergens implicated in cross-reactivity between arthropod species, including house dust mites (HDM), shrimp, and insects. Despite their clinical relevance, a systematic comparison of structural conservation, enzymatic activity, and IgE recognition across allergenic AKs from different taxa is still lacking.

Recombinant AKs from HDM (Der p 20, Der f 20 and like variants), shrimp (Pen m 2), grasshopper (Sch a), crab (Sar s 20), and oyster (Lit v 2) were heterologously expressed and purified. Structural integrity and thermal stability were assessed by circular dichroism spectroscopy. Enzymatic activity was quantified using ATP hydrolysis assays. Sequence and structural comparisons focused on conservation of the catalytic pocket. IgE reactivity was analyzed by ELISA and dot blot assays using sera from shrimp-allergic patients and non-allergic controls.

All recombinant AKs were obtained as soluble, monomeric proteins with comparable secondary structure content and melting temperatures ranging from ~43°C to ~60°C. All variants displayed robust, concentration-dependent ATP hydrolysis, reaching similar maximal activities but differing in apparent activation profiles. Sequence and structural analyses revealed a highly conserved catalytic architecture across species, including key residues involved in substrate binding and catalysis. IgE binding was detected for all AKs tested, with heterogeneous recognition patterns among patients. Notably, preliminary immunological analyses indicated unexpectedly frequent IgE reactivity to insect-derived AKs compared to HDM AKs, prompting ongoing critical re-evaluation of these findings in collaboration with immunology experts.

This study establishes arginine kinases as structurally and functionally conserved pan-allergens across mites, crustaceans, and insects, with preserved catalytic architecture and enzymatic activity despite sequence divergence. The consistent IgE recognition across multiple species confirms the clinical relevance of arginine kinases as drivers of cross-reactivity in arthropod allergy. Differences in IgE reactivity frequencies between allergen sources point to additional immunological factors beyond structural conservation that may shape sensitization patterns. Ongoing expert-driven immunological analyses aim to further resolve these differences and refine their relevance for diagnosis and risk assessment.