The 2S albumin‐like protein from Amaranthus disrupts insect metabolism and triggers antibiosis

BACKGROUND Insect pests cause significant crop losses globally, prompting a need for sustainable control strategies. While chemical pesticides are effective, their environmental impact has increased interest in plant‐derived alternatives. Plants produce defensive proteins, including those that inhibit insect digestive enzymes. Identifying small, stable, and selective inhibitors holds potential for eco‐friendly pest control. RESULTS We report the characterization of a novel 5 kDa protein from Amaranthus hypochondriacus seeds (AhyAI), which effectively inhibits α‐amylases and α‐glucosidases. AhyAI shows high sequence similarity to 2S albumins and exhibits uncompetitive inhibition. Purified AhyAI is stable up to 70 °C and active across acidic to neutral pH. AhyAI inhibited α‐amylases from Spodoptera frugiperda (100%), Oryzaephilus surinamensis (74%), Sitophilus granarius (83%), and Aspergillus oryzae (100%) with half‐maximal inhibitory concentration (IC 50 ) values of 14.4 ± 0.63, 2.12 ± 0.32, 3.82 ± 0.36 and 3.79 ± 0.19 ppm, respectively, but did not inhibit human salivary amylase. It also completely inhibited α‐glucosidases from Spodoptera frugiperda , Sitophilus granarius , and yeast with IC 50 values of 0.93 ± 0.14, 0.29 ± 0.04, and 11.00 ± 0.34 ppm, respectively. In vivo bioassays with Spodoptera frugiperda larvae showed impaired nutrient utilization, growth retardation, and developmental abnormalities in larvae, pupae, and moths upon dietary ingestion of AhyAI. CONCLUSION Disruption of carbohydrate digestion by AhyAI severely affects insect development. Small size, high stability, and pest specificity make AhyAI a promising candidate for eco‐friendly pest control, either in transgenic crops or as part of sprayable formulations.