Biochemical properties of digestive proteases of melon ladybird, Epilachna chrysomelina (Fabricius) (Col.: Coccinellidae)

Document Type : Paper, Persian

Authors

1 Plant Protection Research Department, Mazandaran Agricultural and Natural Resources Research and Education Center, Agricultural Research, Education and Extension Organization (AREEO), Sari, Iran

2 Former student, department of plant protection, faculty of agricultural science, university of guilan

3 Department of Plant Protection, faculty of agricultural science, university of guilan

Abstract

Melon ladybird, Epilachna chrysomelina (Fabricius) (Col.: Coccinellidae)is a major pest of cucurbitaceous plants by feeding on leaves at larval and adult stages. In this study, biochemical properties of digestive proteases of the melon ladybird were studied in its alimentary canal. Enzyme activity in the digestive systems of 2nd, 3rd and 4th larval instars and adults showed that the highest specific enzyme activities occurred in 3rd instar larvae while no significant differences observed in adults. The optimal pH and temperature for protease activity in alimentary canal were found 5 and 30°C, respectively. The inhibitory effect of EDTA, TLCK, TPCK, PMSF and Iodoacetate were determined 44.32, 26.05, 22.27, 19.04 and 18.93% in the gut of E. chrysomelina, respectively. With respect to the highest inhibition rate in proteolytic activity caused by Iodoacetate, cysteine proteinases are considered as the main proteases in the midgut of the melon ladybird. Zymogram pattern in the native gel indicated that the protease enzyme had one isoform in the alimentary canal of the pest.

Keywords

References

Alarcon, F.J., Martınez, T.F., Barranco, P., Cabello, T., Dıaz, M. & Moyano, F.J. (2002) Digestive proteases during development of larvae of red palm weevil, Rhynchophorus ferrugineus (Olivier, 1790) (Coleoptera: Curculionidae). Insect Biochemistry and Molecular Biology 32, 265– 274.
Bird, R. & Hopkin, R. H. (1954) p-Amylolysis: Union of Enzyme and Substrate. Biochemisry Journal 57, 162–165.
Boyd, D. W., Cohen, A. C. & Alverson, D. R. (2002) Digestive enzymes and stylet morphology of Deraeocoris nebulosus (Hemiptera: Miridae), a predacious plant bug. Annals of the Entomological Society of America 95, 395–401.
Bradford, M. (1976) A rapid and sensitive method for quantitation of microgram quantities of protein utilizing the principle of protein–dye binding. Analytical Biochemistry 72, 248–254.

Broadway, R. M., Duffey, S. S., Pearce, G. & Ryan, C. A. (1986) Plant proteinase inhibitors: A defense against herbivorous insects. Entomologia Experimentalis et Applicata 41, 33–38.

Budatha, M., Meur, G. & Datta-Gupta, A. (2008) Identification and characterization of midgut proteases in Achaeta janata and their implication. Biotechnology Letters 30, 305–310.
Chougule, N. P., Doylea, E., Fitchesb E. & Gatehouse, J. A. (2008) Biochemical characterization of midgut digestive proteases from Mamestra brassicae (cabbage moth; Lepidoptera: Noctuidae) and effect of soybean Kunitz inhibitor (SKTI) in feeding assays. Journal of Insect Physiology 54, 563–572.
Christopher, M. S. M. & Mathavan, S. (1985) Regulation of digestive enzyme activity in the larvae of Catopsilia crocale. Journal of Insect Physiology 31, 217–221.
Davis, B. J. (1964) Disc electrophoresis II. Method and application to human serum proteins. Annals of the New York Academy of Sciences 12, 404–427.
Dow J. A. T. (1986) Insect midgut function. Advances in Insect Physiology 19, 187–329.
Erlanger, B., Kokowsky N. & Cohen, W. (1961) The preparation and properties of two new chromogenic substrates of trypsin. Archive Biochemistry and Biophysics 95, 271–278.
Esmaili, M., Mirkarimi, A. & Azmayeshfard, P. (2004) Agricaltural entomology. 500 pp. Tehran University Press. [In Persian].
Fabrick, J., Behnke, C., Czapla, T., Bala, K., Rao, A.G., Kramer, K.J. & Reeck, G.R. (2002) Effects of a potato cysteine proteinase inhibitor on midgut proteolytic enzyme activity and growth of the southern cornrootworm, Diabrotica undecimpunctata howardi (Coleoptera: Chrysomelidae). Insect Biochemistry and Molecular Biology 32, 405–415.
Garcia-Carreno, F. L., Dimes, L. E. & Haard, N .F. (1993) Substrate-gel electrophoresis for composition and molecular weight of proteinases orproteinaceous protease inhibitors. Analytical Biochemistry 214, 61–69.
Hummel, B. C. W. (1959) A modified spectrophotometric determinations of chymotrypsin, trypsin and thrombin. Canadian Journal of Biochemistry and Physiology 37, 1393–1399.
Izadpanah, K. (1989) Characterization of squash mosaic virus. Journal of Plant Pathology 25, 1–4. [In Persian].
Kuroda, M., Ishimoto, M., Suzuki, K., Kondo, H., Abe, K., Kitamura, K. & Arai, S. (1996) Oryzacystatins exhibits growth-inhibitory and lethal effects on different species of bean insects pest, Callosobruchus chinensis (Coleoptera) and Riptortus clavatus (Hemiptera). Bioscience, Biotechnology and Biochemistry 60, 209–212.
Lemos, F. J. A. & Terra, W. R. (1991) Properties and interacellular distribution of cathepsin D-like proteinase active at the acid region of Musca domestica midgut. Insect Biochemistry 21, 457–412.
Lemos, F. J. A., Campos, F. A. P., Silva, C. P. & Xavier-Filho, J. (1990) Proteinases and amylases of larval midgut of Zabrotes subfasciatus reared on cowpea (Vigna unguiculata) seeds. Entomologia Experimentalis et Applicata 56, 219–227. 
Leo, F. D., Volpicella, M., Licciulli, F., Liuni, S., Gallerani, R. & Ceci, L. R. (2002) PLANT-PIs: a database for plant protease inhibitors and their genes. Nucleic Acids Research 30, 347–348.
Montesdeoca, M., Lobo, M. G., Casañas, N., Carnero, A., Castañera P. & Ortego, F. (2005) Partial characterization of the proteolytic enzymes in the gut of the banana weevil, Cosmopolites sordidus, and effects of soybean Kunitz trypsin inhibitor on larval performance. Entomologia Experimentalis et Applicata 116, 227–236.
Novillo, C., Castanera, P. & Ortego, F. (1997) Characterization and distribution of chymotrypsin-like and other digestive proteases in Colorado potato beetle larvae. Archive of Insect Biochemistry and Physiology 36, 181–201.
Oppert, B., Morgan, T. D., Hartzer, K., Lenarcic, B., Galesa, K., Brzin, J., Turk, V., Yoza, K., Ohtsubo, K. & Kramer, K. J. (2003) Effects of proteinase inhibitors on digestive proteinases and growth of the red flour beetle, Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae). Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology 134, 481–490.
Patankar, A. G., Giri, A. P., Harsulkar, A. M., Sainani, M. N., Deshpande, V. V., Ranjekar, P. K. & Gupta, V. S. (2001) Complexity in specificities and expression of Helicoverpa armigera gut proteinases explains polyphagous nature of the insect pest. Insect Biochemistry and Molecular Biology 31, 453–464.
Ryan, C. A. (1999) Protease inhibitors in plants: genes for improving defenses against insects and pathogens. Annual Review of Phytopathology 28, 425–449.
Shaheen, A. H., Elezz, A. A. &Assem, M. A. (1973) Chemical control of cucurbits pests at Komombo. Agricultural Research Review 51 (1), 103–107.
Tatli, A., Bandani, A. & Naghdi, M. (2010) Study of the digestive protease in the elm leaf beetle Xanthogaleruca luteola (Col.,: Chrysomylidae). Proceedings of the 19th Iranian Plant Protection Congress, 9-12 Aug., Tehran, Iran, p. 300. [In Persian].
Terra, W. R. & Cristofoletti, P. T. (1996) Midgut proteases in three divergent species of Coleoptera. Comparative Physiology and Biochemistry 113B, 725–730.
Zahedi, K. (1993) Vegetable and ornamental plants pests in Iran and methods of control. University Press. 143pp. [In Persian].
Zang, F., Zhu, Y. & Cohen, A. C. (2002). Molecular cloning and partial characterization of a trypsin-like protein in salivary glands of Lygus Hesperus (Hemiptera: Miridae). Insect Biochemistry and Molecular Biology 32, 455–464.
Zhu-Salzman, K.,  Koiwa, H., Salzman, R. A., Shade R. E. &  Ahn, J. E. (2003) Cowpea bruchid Callosobruchus maculatus uses a three-component strategy to overcome a plant defensive cysteine protease inhibitor. Insect Molecular Biology 12, 135–145.

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