1H, 15N and 13C backbone and side-chain resonance assignments of Amblyomin-X Kunitz domain

Amblyomin-X is a Kunitz-type inhibitor of factor Xa (FXa) from the tick Amblyomma sculptum with promising anticoagulant activity and potential therapeutic applications. Despite the relevance, the molecular basis of its interaction with FXa remains poorly understood, limiting a detailed understanding of its mechanism of action. In this study, we report the 1 H, 13 C, and 15 N resonance assignments of the Amblyomin-X Kunitz domain obtained by multidimensional NMR spectroscopy. High-quality spectra enabled extensive assignment, reaching 97.9% completeness for backbone assignments and 79.8% for side-chain assignments. The 1 H- 15 N HSQC spectrum shows excellent signal dispersion, consistent with a well-folded protein in solution. Furthermore, chemical shift-based structure analysis reveals elements that agree with the canonical Kunitz fold, including characteristic β-strands and helical regions. These results provide the first detailed NMR characterization of the Amblyomin-X Kunitz domain l. The resonance assignments presented here constitute a critical foundation for future structural and dynamical studies, including analyses of protein-ligand interactions. Ultimately, this work contributes to a deeper understanding of the molecular determinants governing FXa inhibition by Amblyomin-X and supports ongoing efforts to develop novel anticoagulant strategies based on Kunitz-type inhibitors.