Eur. J. Entomol. 102 (2): 147-153, 2005 | DOI: 10.14411/eje.2005.023

Trypsin-like activity of membrane-bound midgut proteases from Anticarsia gemmatalis (Lepidoptera: Noctuidae)

Luciana Pereira XAVIER1, Maria Goreti ALMEIDA OLIVEIRA*,1,2, Raul Narciso Carvalho GUEDES2, Agenor Valarades SANTOS3, Salvatore Giovanni DE SIMONE4
1 Departament of Biochemistry and Molecular Biology, Institute of Biotechnology Applied to Agriculture and Animal Science (BIOAGRO), Federal University of Viçosa, Viçosa, MG 36571-000, Brazil
2 Departament of Animal Biology, Federal University of Viçosa, Viçosa, MG 36571-000, Brazil
3 Departament of Microbiology, Institute of Biotechnology Applied to Agriculture and Animal Science (BIOAGRO), Federal University of Viçosa, Viçosa, MG 36571-000, Brazil
4 Departament of Biochemistry and Molecular Biology, Foundation Oswaldo Cruz Institute (FIOCRUZ), Rio de Janeiro, RJ 21045-900, Brazil

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.

Keywords: Trypsin, proteases, velvetbean caterpillar, Lepidoptera, Noctuidae, Anticarsia

Received: July 1, 2004; Revised: November 29, 2004; Accepted: November 29, 2004; Published: May 3, 2005  Show citation

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XAVIER, L.P., ALMEIDA OLIVEIRA, M.G., GUEDES, R.N.C., SANTOS, A.V., & DE SIMONE, S.G. (2005). Trypsin-like activity of membrane-bound midgut proteases from Anticarsia gemmatalis (Lepidoptera: Noctuidae). EJE, 102(2), 147-153. doi: 10.14411/eje.2005.023
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