Cystatins are cysteine protease inhibitors involved in defence mechanisms against pests and pathogens. Here, the cystatin CaCPI gene was isolated from a cDNA library of the Siam tulip (Curcuma alismatifolia cv. Chiang Mai Pink was then transformed into Escherichia coli strain BL21-Star to produce a recombinant CaCPI protein. After induction with 1 mM IPTG, the cell lysate of E. coli-carrying pDEST17-CaCPI generated a CaCPI protein about 12 kDa in size as measured using SDS-PAGE. Pre-incubation of the 5-30 µM CaCPI protein samples with 5 µM papain is known to decrease papain activity. Antifungal activities of the purified recombinant CaCPI protein against phytopathogenic fungi were tested. The CaCPI protein could suppress mycelium growth of Fusarium oxysporum, Colletotrichum capsici, and Pyricularia grisea phytopathogenic fungi.
Cystatin is a cysteine protease inhibitor that can regulate the proteolytic process of cysteine proteases by binding to the active site of those target enzymes. Plant cystatin has several reported roles, including in: protein turnover during development; programmed cell death; and plant defense mechanisms against insects, nematodes and phytopathogenic fungi. In this experiment, the CaCPI gene was isolated from the cDNA library of the Siam tulip (Curcuma alismatifolia cv. Chiang Mai Pink). Its full length cDNA sequence is 601 base pair, containing 372 base pair in an open reading frame encoding 123 amino acids and 32 and 197 base pair in the 5'and 3' untranslated regions, respectively. The deduced amino acid sequence consists of a putative N-terminal secretory signal peptide of 22 amino acids and an estimated molecular mass for the mature protein of 11.239 kDa. The CaCPI protein contains all of the highly conserved blocks, including Gly 31-Gly 32 , the reactive site motif QXVXG (Q 76 V 77 V 78 A 79 G 80), P 106-W 107 , and the LGRFAVDQHN block that are common among plant cystatins. The CaCPI gene was cloned into a pDEST17 expression vector and then transformed into the Escherichia coli strain BL21-Star for recombinant CaCPI protein production. After induction with 1 mM IPTG, the cell lysate of E. coli carrying pDEST17-CaCPI generated a CaCPI protein about 12 Kda in size on SDS-PAGE. This cell lysate inhibited papain activity.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.