Characterization and partial purification of the digestive proteases of the black field cricket, Teleogryllus commodus (Walker): Elastase is a major component

Midgut proteinase activities of three keratinolytic larvae, Hofmannophila pseudospretella, Tineola bisselliella, and Anthrenocerus australis, and the effect of proteinase inhibitors on proteolysis

“…After centrifugation, the supernatant extracts were analysed immediately. The proteinase substrate, "k-casein, was synthesized [24] and assayed [191 as described previously by measuring trichloroacetic acid-soluble peptides in a Packard 1500 liquid scintillation analyser. Specific amidase activities were assayed [24] by measuring the rate of p-nitroanilide production at 410 nm in a Dynateck MR5000 ELISA plate reader as follows: trypsin-like activity with 1 mM N-benzoyl-DL-arginine-p-nitroanilide, chymotrypsin-like activity with 1.25 mM N-benzoyl-L-tyrosine-p-nitroanilide, SAAPLpNA* hydrolyzing activity with 0.5 mM N-succinyl-L-alanyl-L-alany-L-prolyl-L-leu~~p-~troanilide~ and aminopeptidase activity with 0.5 mM L-leucine-p-nitroanilide.…”

“…The proteinase substrate, "k-casein, was synthesized [24] and assayed [191 as described previously by measuring trichloroacetic acid-soluble peptides in a Packard 1500 liquid scintillation analyser. Specific amidase activities were assayed [24] by measuring the rate of p-nitroanilide production at 410 nm in a Dynateck MR5000 ELISA plate reader as follows: trypsin-like activity with 1 mM N-benzoyl-DL-arginine-p-nitroanilide, chymotrypsin-like activity with 1.25 mM N-benzoyl-L-tyrosine-p-nitroanilide, SAAPLpNA* hydrolyzing activity with 0.5 mM N-succinyl-L-alanyl-L-alany-L-prolyl-L-leu~~p-~troanilide~ and aminopeptidase activity with 0.5 mM L-leucine-p-nitroanilide. Following initial investigation, determinations of enzyme levels and inhibition studies were carried out at a pH reflecting both the optimum for proteolysis and within the range of observed pH for the central midgut [10,11], i.e., at pH 10.0 in 50 mM CAPS-NaOH buffer for the two lepidopteran species and at pH 8.0 in 0.1 M Tris-HC1 buffer for the coleopteran species.…”

A Plant Chloroplast Glutamyl Proteinase

“…In inhibition studies the inhibitor and enzyme were incubated together for at least 10 min before the substrate was added to ensure that equilibrium between the proteinase and the inhibitor was established. At least six time points were read in any assay with the plate incubated at 30°C between readings, and the results were collected by a computer and analyzed using programs similar to those described by Christeller et al (1990). Only those points that showed a linear time course were used to calculate the reaction rate.…”

Protein and cDNA sequences of Bowman-Birk protease inhibitors from the cowpea (Vigna unguiculata Walp.)

“…The serine proteases of the majority of lepidopteran pests of major economic importance which have been determined are trypsin-like rather than chymotrypsin-like [ 1 ]. Chymotrypsinlike enzymes are, however, present in the guts of some insects, and the recent identification of elastase-like enzymes as major proteases in a number of herbivorous insects [7] no doubt accounts for the (previously puzzling) presence of this class of inhibitors in plants. The mixture of inhibitors in a particular plant must reflect the co-evolutionary processes in particular plant/ insect complexes established before agricultural usage.…”