In the present report we study the proteolytic activity of the excretion-secretion and crude extracts of different stages of Trichinella spiralis (Owen, 1835) Railliet, 1895, (muscle-stage larvae, adult worms before and after mating, and newborn larvae) using natural substrates (structural and hematic mammalian proteins). The analysis of the results allow us to set up a certain stage-specificity, as well as an important relationship between the protease patterns throughout the parasite life cycle and how the parasite may overcome both mechanical and humoral barriers within the host. Muscle-stage larvae present a great activity against structural proteins (collagen), while newborn larvae and adult worms degrade principally hematic proteins (hemoglobin, fibrinogen and immunoglobulin G).
Membrane-bound proteases from preparations of the midgut of 5th instar velvetbean caterpillars, Anticarsia gemmatalis (Hübner) were obtained by resuspension of the pellet obtained after 100,000 g centrifugation. As expected of trypsin-like proteases, they hydrolyzed casein and the synthetic substrates N-α-benzoyl-L-Arg-p-nitroanilidine (L-BApNA) and N-α-p-tosyl-L-Arg methyl ester (L-TAME). Higher activities were observed at 50°C, and at pH 8.5 and 8.0 for both synthetic substrates L-BApNA and L-TAME. The membrane-bound proteases were inhibited by EDTA, phenylmethan sulphonyl fluoride (PMSF), tosyl-L-lysine chloromethyl ketone (TLCK), benzamidine and aprotinin. TLCK and benzamidine were particularly active inhibitors. The KM-values obtained were 0.23 mM for L-BApNA and 92.5 µM for L-TAME. These results provide evidence for the presence of membrane-bound trypsin-like proteases in the midgut of the velvetbean caterpillar, a key soybean pest in warm climates. The interaction between A. gemmatalis digestive proteases and soybean protease inhibitors has potentially important consequences for soybean breeding programs.