Proteolytic activation of canatoxin, a plant toxic protein, by insect cathepsin‐like enzymes

Abstract

Canatoxin is a protein isolated from jackbean (Canavalia ensiformis), seeds. Injected intraperitoneally, the toxin is lethal to mice but it is inactive if given orally. Canatoxin is also lethal when fed to insects with cathepsin-based digestion while insects with trypsin-based digestion are not affected. The hypothesis that canatoxin is proteolytically activated by cathepsins was investigated. Experiments were performed with 4(th) instar and adult Rhodnius prolixus fed meals containing canatoxin (2.5 microg/mg weight body). While 100% of nymphs died, no effect was observed in adults. Hemolymph taken from nymphs and adults showed the presence of canatoxin's proteolytic fragments. Reduced lethality was seen in R. prolixus 4(th) instars fed meals containing canatoxin and inhibitors of cathepsin enzymes, E-64 (2.0 microM) or Pepstatin-A (2. 0 microM). In another approach, canatoxin was digested in vitro with enzymes from the bruchid, Callosobruchus maculatus, and the resulting peptides were tested in R. prolixus. Three groups of toxic peptides (8,000-12,000 kD range) were separated by gel-filtration. When these peptides were fed to the insects simultaneously with the cathepsin inhibitors, no protective effect was seen. These results confirm the proteolytic activation of canatoxin by insect cathepsin-like enzymes to produce entomotoxic peptide(s). Furthermore, our data point towards overlooked differences in the digestive physiology of distinct life stages of R. prolixus. Arch.

Jasmonate-inducible plant enzymes degrade essential amino acids in the herbivore midgut

“…This proposed defensive role for tomato TD is consistent with the enzyme's extraordinarily high expression in f lowers (29) and JA-induced leaves (16,40). Increasing evidence of proleolytic activation of defensive proteins in the insect gut (41,42) indicates that this phenomenon may be more common than previously realized. The idea that plant enzymes can exert antinutritional effects on phytophagous insects by perturbing amino acid homeostasis in the digestive tract may be extended to other classes of plantderived nutrients, including lipids, carbohydrates, and vitamins.…”

Chagasin, the endogenous cysteine-protease inhibitor ofTrypanosoma cruzi, modulates parasite differentiation and invasion of mammalian cells

“…The biological implications of secreted chagasin-cruzipain complexes are not clear. Epimastigotes replicate in the gut of the insect vector, a protease-rich environment (Garcia et al, 1978;Ferreira-da Silva et al, 2000). In this scenario, it is not obvious how chagasin would dissociate from cruzipain, thus becoming available to inhibit the vector-derived CPs.…”

Pliable natural biocide: Jaburetox is an intrinsically disordered insecticidal and fungicidal polypeptide derived from jack bean urease

“…This toxicity involves an internal polypeptide of 10 kDa (pepcanatox), released from the protein upon hydrolysis by cathepsin-like digestive enzymes of insects such as Callosobruchus maculatus, Rhodnius prolixus, Nezara viridula, Dysdercus peruvianus and Oncopeltus fasciatus [13][14][15][16] (Fig. 1A,B).…”

“…The only unassigned residues were E49 and K62. Nearly complete assignments were achieved for the other backbone nuclei (100% for 13 Ca, 100% for Secondary structure propensity of Jaburetox from NMR chemical shifts Disordered proteins tend to feature only few longrange tertiary contacts, and therefore local structural constraints on the backbone are important in order to describe the ensemble of possible conformers of the protein. In this perspective, the structural information encoded in the chemical shifts of backbone nuclei has emerged as a key indicator of IDP ensemble properties [48].…”

“…Calculation of the structure of Jaburetox using NMR spectroscopy NOE cross-peaks were identified, assigned and integrated in 15 N-edited and 13 C-edited 3D NOESY-HSQC spectra TD is the total number of real+imaginary points collected. SI is the total number of real+imaginary points before the Fourier transform.…”