RT Journal Article
T1 Analysis of Protein Inhibitors of Trypsin in Quinoa, Amaranth and Lupine Seeds. Selection and Deep Structure–Function Characterization of the Amaranthus caudatus Species
A1 Hernández de la Torre, Martha
A1 Covaleda Cortés, Giovanni
A1 Montesinos, Laura
A1 Covaleda, Daniela
A1 Ortiz, Juan C.
A1 Piñol, Jaume
A1 Bautista Santa Cruz, José Manuel
A1 Castillo, J. Patricio
A1 Reverter, David
A1 Avilés, Francesc Xavier
AB Protease inhibitors are biomolecules with growing biotechnological and biomedical relevance, including those derived from plants. This study investigated strong trypsin inhibitors in quinoa, amaranth, and lupine seeds, plant grains traditionally used in Andean South America. Amaranth seeds displayed the highest trypsin inhibitory activity, despite having the lowest content of aqueous soluble and thermostable protein material. This activity, directly identified by enzymatic assay, HPLC, intensity-fading mass spectrometry (IF-MS), and MS/MS, was attributed to a single protein of 7889.1 Da, identified as identical in Amaranthus caudatus and A. hybridus, with a Ki of 1.2 nM for the canonical bovine trypsin. This form of the inhibitor, which is highly homogeneous and scalable, was selected, purified, and structurally–functionally characterized due to the high nutritional quality of amaranth seeds as well as its promising agriculture–biotech–biomed applicability. The protein was crystallized in complex with bovine trypsin, and its 3D crystal structure resolved at 2.85 Å, revealing a substrate-like transition state interaction. This verified its classification within the potato I inhibitor family. It also evidenced that the single disulfide bond of the inhibitor constrains its binding loop, which is a key feature. Cell culture assays showed that the inhibitor did not affect the growth of distinct plant microbial pathogen models, including diverse bacteria, fungi, and parasite models, such as Mycoplasma genitalium and Plasmodium falciparum. These findings disfavour the notion that the inhibitor plays an antimicrobial role, favouring its potential as an agricultural insect deterrent and prompting a redirection of its functional research
PB MDPI
SN 1661-6596
YR 2025
FD 2025
LK https://hdl.handle.net/20.500.14352/118422
UL https://hdl.handle.net/20.500.14352/118422
LA eng
NO Hernández de la Torre, M., Covaleda-Cortés, G., Montesinos, L., Covaleda, D., Ortiz, J. C., Piñol, J., Bautista, J. M., Castillo, J. P., Reverter, D., & Avilés, F. X. (2025). Analysis of Protein Inhibitors of Trypsin in Quinoa, Amaranth and Lupine Seeds. Selection and Deep Structure–Function Characterization of the Amaranthus caudatus Species. International Journal of Molecular Sciences, 26(3), 1150. https://doi.org/10.3390/ijms26031150
NO Author Contributions: Conceptualization: M.H.d.l.T., G.C.-C., J.P.C., D.R. and F.X.A.; experimental acquisition, data analysis, writing, and editing: M.H.d.l.T., G.C.-C., L.M., D.C., J.C.O., J.P., J.M.B., J.P.C., D.R. and F.X.A.; general writing and revision: M.H.d.l.T., G.C.-C., J.P., J.M.B., D.R. and F.X.A.; corresponding author: F.X.A. All authors have read and agreed to the published version of the manuscript
NO Ministerio de Ciencia, Innovación y Universidades (España)
NO Generalitat de Catalunya. ICREA
DS Docta Complutense
RD 10 abr 2025