Characterisation of cysteine proteinases responsible for digestive proteolysis in guts of larval western corn rootworm (Diabrotica virgifera) by expression in the yeast Pichia pastoris

Deleterious effects of plant cystatins against the banana weevil Cosmopolites sordidus

“…By contrast, the E-64 derivative CA-074, specific to cathepsin B-like enzymes (Murata et al, 1991), showed no inhibitory activity against most Z-RR-MCA-and Z-FR-MCA-hydrolyzing enzymes (Fig. 3B), as also reported earlier for midgut cysteine cathepsins of the Western corn rootworm Diabrotica virgifera (Bown et al, 2004).…”

“…By contrast, the E-64 derivative CA-074, specific to cathepsin B-like enzymes (Murata et al, 1991), showed no inhibitory activity against most Z-RR-MCA-and Z-FR-MCA-hydrolyzing enzymes (Fig. 3B), as also reported earlier for midgut cysteine cathepsins of the Western corn rootworm Diabrotica virgifera (Bown et al, 2004).Unlike cysteine-type inhibitors (not shown), the potent trypsin inhibitor bovine aprotinin inhibited most trypsin-like and some chymotrypsin-like activities in the extracts, compared to the soybean Bowman-Birk (trypsin-chymotrypsin) inhibitor completely inactivating both Z-R-MCA-and suc-AAPF-MCA-hydrolyzing proteinases (Fig. 3C).…”

“…3A), suggesting the presence of cathepsin L-like, cathepsin B-like and aminopeptidase (cathepsin H-like) activities in larval extracts, as observed earlier for a number of coleopteran insects (e.g. Thie and Houseman, 1990;Michaud et al, 1993;Bown et al, 2004;Montesdeoca et al, 2005; Vinokurov et al, 2006a,b;Prabhakar et al, 2007).…”

“…3A), suggesting the presence of cathepsin L-like, cathepsin B-like and aminopeptidase (cathepsin H-like) activities in larval extracts, as observed earlier for a number of coleopteran insects (e.g. Thie and Houseman, 1990;Michaud et al, 1993;Bown et al, 2004;Montesdeoca et al, 2005; Vinokurov et al, 2006a,b;Prabhakar et al, 2007).Trypsin-and chymotrypsin-like serine proteases hydrolyzing, respectively, Z-R-MCA and succinyl-AAPF-MCA were detected at pH 8.5 (Fig. 3A) but merely detectable at pH 6.5 (not shown), which points again to the presence of serine proteinases in the midgut extracts, as also reported for other Coleoptera predominantly relying on cysteine proteases for dietary protein digestion (e.g.…”

“…3B). Overall, the well-documented preference of coleopteran cathepsin L-like enzymes for Z-FR-MCA despite the Z-RR-MCA-hydrolyzing activity of some isoforms (Bown et al, 2004;Cristofoletti et al, 2005), the specificity of cathepsin B-like enzymes for Z-RR-MCA (Mort, 1998), and the differential inhibitory effects of Z-FY-DMK and human stefin A against banana weevil Z-FR-and Z-RR-hydrolyzing proteinases (Student t-test; P<0.01) (Fig. 3B) suggest the occurrence of cathepsin L-like and cathepsin B-like proteinase populations in the banana weevil midgut, both sensitive to E-64 and cystatins but exhibiting distinct preferences for the Z-FR-MCA and Z-RR-MCA substrates.…”