Multifunctional amaranth cystatin inhibits endogenous and digestive insect cysteine endopeptidases: A potential tool to prevent proteolysis and for the control of insect pests

Abstract In a previous study, the amaranth cystatin was characterized. This cystatin is believed to provide protection from abiotic stress because its transcription is induced in response to heat, drought, and salinity. It has also been shown that recombinant amaranth cystatin inhibits bromelain, ficin, and cysteine endopeptidases from fungal sources and also inhibits the growth of phytopathogenic fungi. In the present study, evidence is presented regarding the potential function of amaranth cystatin as a regulator of endogenous proteinases and insect digestive proteinases. During amaranth germination and seedling growth, different proteolytic profiles were observed at different pH levels in gelatin‐containing SDS‐PAGE. Most of the proteolytic enzymes detected at pH 4.5 were mainly inhibited by trans ­epoxysuccinyl‐leucyl amido(4‐guanidino)butane (E‐64) and the purified recombinant amaranth cystatin. Furthermore, the recombinant amaranth cystatin was active against insect proteinases. In particular, the E‐64‐sensitive proteolytic digestive enzymes from Callosobruchus maculatus , Zabrotes subfasciatus , and Acanthoscelides obtectus were inhibited by the amaranth cystatin. Taken together, these results suggest multiple roles for cystatin in amaranth, specifically during germination and seedling growth and in the protection of A. hypochondriacus against insect predation. Amaranth cystatin represents a promising tool for diverse applications in the control of insect pest and for preventing undesirable proteolytic activity.