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The coffee berry borer, Hypothenemus hampei, is one of the most devastating coffee pests (Coffea arabica). Aspartic proteases were identified in protein extracts of adult insects, showing two bands of proteolytic activity in zymograms and a maximum activity at pH 2.5 in the spectrophotometric assays. Considering its inhibition by pepstatin A and hemoglobin hydrolysis this enzyme was classified as cathepsin D. Seeds extracts of Lupinus bogotensis, Brachiaria humidicola, Amaranthus hypochondriacus, Phaseolus acutifolius, Phaseolus coccineus, Hyptis suaveolens, Centrosema pubescens, and Trifolium repens were evaluated to identify aspartic protease inhibitors. The greatest inhibitory activity of aspartic proteases was found in L. bogotensis extract with a specific activity of 74.1 IU/mg, compared with extracts of H. suaveolens, B. humidicola, and A. hypochondriacus that showed a lower inhibition of aspartic proteases. Other protein extracts did not inhibit aspartic protease activity. The proteolytic activity of H. hampei was inhibited by 90% with 100 g of crude extract of L. bogotensis, whereas it took 1 mg of crude extract of H. suaveolens and B. humidicola to inhibit the activity at 70 and 60% respectively. The zymograms identified potential protease inhibitors in extracts of L. bogotensis, H. suaveolens, B. humidicola and A. hypochondriacus that blocked in vitro activity of aspartic proteases of the insect. The expression of protease inhibitor genes in coffee is an alternative to obtain resistant varieties.

MOLINA-V, D., BLANCO-LABRA, A., & ZAMORA-E, H. (2011). Plant protease inhibitors effective against aspartic proteases from Coffee Berry Borer (Hypothenemus hampei). Revista Colombiana De Entomología, 37(2), 183–191. https://doi.org/10.25100/socolen.v37i2.9072

AGUIRRE, C.; VÁLDES, S.; MENDOZA, G.; ROJO, A.; BLANCO, A. 2004. A novel 8.7 kDa protease inhibitor from chan seeds (Hyptis suaveolens L.) inhibits proteases from larger grain borer Prostephanus truncates (Coleoptera: Bostrichidae). Comparative Biochemistry and Physiology. Part B: Biochemistry & Molecular Biology 138. 81-89.

AHN, J. E.; ZHU-SALZMAN, K. 2009. CmCatD, a catepsin D-like protease has a potential role in insect defense against a phytocystatin. Journal of Insect Physiology 55: 678-685.

AMIRHUSIN, B.; SHADE, R. E.; KOIWA, H.; HASEGAWA, P. M.; BRESSAN, R. A.; MURDOCK, L.; ZHU-SALZMAN, K. 2007. Protease inhibitors from several classes work synergistically against Callosobruchus maculatus. Journal of Insect Physiology 53. 734-740.

BARBOSA, A. E.; ALBUQUERQUE, E.; SILVA, M.; SOUZA, D.; OLIVEIRA-NETO, O.; VALENCIA, A.; ROCHA, T.; GROSSI-DE-SA, M. F. 2010. -amylase inhibitor1 gene from Phaseolus vulgaris expressed in Coffea Arabica plants inhibits -amylases from coffee berry borer. BMC Biotechnology 10:44.

BERGAMIN, J. 1943. Contribuição para o conhecimento da biologia da broca do café Hypothenemus hampei (Ferrari, 1867) (ColIpidae). Arquivos do Instituto Biologico 14: 31-72.

BLANCO, A.; MARTÍNEZ, N.; SANDOVAL, L.; DELANO, J. 1996. Purification and characterization of a digestive Cathepsin D proteinase isolated from Tribolium castaneum larvae (Herbst). Insect Biochemistry and Molecular Biology 26: 95-100.

BOLDBAATAR, D.; SIKASUNGE, C. S.; BATTSETSEG, B.; XUAN, X.; FUJISAKI, K. 2006. Molecular cloning and functional characterization of an aspartic protease from the hard tick Haemaphysalis longicornis. Insect Biochemistry and Molecular Biology 36: 25-36.

BUSTILLO, A. E. 2008. Aspectos sobre la broca del café, Hypothenemus hampei, en Colombia. P. 388-418. En: Alex Enrique Bustillo (ed.). Los insectos y su manejo en la caficultura colombiana. FNC. Chinchiná, Colombia. 466 p.

CHO, W. L.; RAICKEL, A. S. 1992. Cloning of cDNA for mosquito lysosomal aspartic protease. Sequence analysis of an insect lysosomal enzyme similar to cathepsins D and E. The Journal of Biological Chemistry 267: 21823-21829.

CHRISTELLER, J. T.; FARLEY, P. C.; RAMSAY, R. J.; SULLIVAN, P. A.; LAING, W. A. 1998. Purification, characterization and cloning of an aspartic proteinase inhibitor from squash phloem exudate. European Journal of Biochemistry 254: 160-167.

DUQUE, H.; BAKER, P. 2003. Case studies in CBB economics. p. 51-63. En: Devouring Profit. The socio economics of coffee berry borer IPM. The Commodities Press-CABI-CENICAFE (ed.). Chinchiná, Colombia. 106 p.

FRANCO, O. L.; DIAS, S. C.; MAGALHAES C. P.; MONTEIRO, A. C.; BLOCH, Jr. C.; MELO, F. R.; OLIVEIRA-NETO, O. B.; MONNERAT, R. G.; GROSSI-DE-SA, M. F. 2004. Effects of soybean Kunitz trypsin inhibitor on the cotton boll weevil (Anthonomus grandis). Phytochemistry 65: 81-89.

GALLARDO, F.; ARAYA, H.; PAK, N.; TAGLE, M. A. 1974. Toxic factors in chilean legumes II. Trypsin inhibitor activity. Archivos Latinoamericanos de Nutrición 24: 183-189.

GATEHOUSE, J. A. 2002. Plant resistance towards insect herbivores: a dynamic interaction. New Phytologist 156: 145-169.

GIRI, A. P.; HARSULKAR, A. M.; KU, M. S. B.; GUPTA, V. S.; DESHPANDE, V. V.; RANJEKAR, P. K.; FRANCESCHI, V. R. 2003. Identification of potent inhibitors of Helicoverpa armigera gut proteinase from winged bean seeds. Phytochemistry 63: 523-532.

HARSULKAR, A. M.; GIRI, A. P.; PATANKAR, A. G.; GUPTA, V. S.; SAINANI, M. N.; RANJEKAR, P. K.; DESHPANDE, V. V. 1999. Succesive use of non-host plant proteinase inhibitors required for effective inhibition of Helicoverpa armigera gut proteinases and larval growth. Plant Physiology 121: 497-506.

HOUSEMAN, J. G.; DOWNE, A. E. R. 1983. Cathepsin D-like activity in the posterior midgut of Hemipteran insects. Comparative Biochemistry and Physiology. Part B: Biochemistry & Molecular Biology 75: 509-512.

JARAMILLO, J.; BORGEMEISTER, C.; BAKER, P. 2006. Coffee berry borer Hypothenemus hampei (Coleoptera: Curculionidae): Searching for sustainable control strategies. Bulletin of Entomological Research 96: 1-12.

JONGSMA, M. A.; BOLTER, C. J. 1997. The adaptation of insects to plant proteinase inhibitors. Journal of Insect Physiology 43: 885-896.

JOHNSON, R.; NARVAEZ, J.; GYNHEUNG, A.; RYAN, C. 1989. Expression of proteinase inhibitors I and II in transgenic tobacco plants: Effects on natural defense against Manduca sexta larvae. Proceedings of the National Academy of Sciences of the United States of America 86: 9871-9875.

LASKOWSKI, M. Jr.; QASIM, M. A.; LU, S. M. 2003. Interaction of standard mechanism, canonical protein inhibitors with serine proteinases. In Protein-Protein Recognition (Kleanthous, C., ed.), pp. 228-279, Oxford University Press, Oxford.

LEE, K. S.; KIM, B. Y.; CHOO, Y. M.; YOON, H. J.; KANG, P. D.; WOO, S. D.; SOHN, H. D.; ROH, J. Y.; GUI, Z. Z.; JE, Y. H.; JIN, B. R. 2009. Expresión profile of cathepsin D in the fat body of Bombix mori during metamorphosis. Comparative Biochemistry and Physiology. Part B: Biochemistry & Molecular Biology 154: 188-194.

LEMOS, K. J. A.; CAMPOS, F. A. P.; SILVA, C. P.; XAVIERFILHO, J. 1990. Proteinases and amylases of larval midgut of Zabrotes subfasciatus on cowpea (Vigna unguiculata) seeds. Entomologia Experimentalis et Applicata 56: 21.

LENNEY, J. F. 1975. Three yeast proteins that specifically inhibit yeast proteases A, B and C. The Journal of Bacteriology 122: 1265-1273.

LI, X.; SCHULER, M, A.; BERENBAUM, M. R. 2002. Jasmonate and salicylate induce expresión of herbivore cytochrome P450 genes. Nature 419: 712-715.

LIU, Y. L.; SALZMAN, R. A.; PANKIW, T.; ZHU-SALZMAN, K. 2004. Transcriptional regulation in southern corn rootworm larvae challenged by soyacystatin N. Insect Biochemistry and Molecular Biology 34, 1069-1077.

MACEDO, M. L.; FREIRE, M. G.; SILVA, M. B.; COELHO, M. B. 2007. Insecticidal action of Bauhinia monandra leaf lectin (BmoLL) against Anagasta kuehniella (Lepidoptera: Pyralidae); Zabrotes subfasciatus and Callosobruchus maculatus (Coleóptera: Bruchidae). Comparative Biochemistry and Physiology. Part A: Molecular & Integrative Physiology 146: 486-498.

MADR-IICA-OAC (Ministerio de Agricultura y Desarrollo Rural, Instituto Interamericano de Cooperación en la Agricultura, y Observatorio de Agrocadenas de Colombia). 2008. Información estadística. <http://www.agrocadenas.gov.co/home.htm> (Fecha última revisión: 20 junio 2010. Fecha último acceso: [10 julio 2010].

MARES, M.; MELOUN, B.; PAVLIK, M.; KOSTKA, V.; BAUDYS, M. 1989. Primary structure of the cathepsin D inhibitor from potatoes and its structural relationship to soybean trypsin inhibitor family. FEBS Letters 251. 94-98.

MENDIOLA, E.; VALENCIA, A.; VALDÉS, S.; DÉLANO, J.; BLANCO, A. 2000. Digestive amylase from the larger grain borer, Prostephanus truncates Horn. Comparative Biochemistry and Physiology. Part B: Biochemistry & Molecular Biology

: 425-433.

MOLINA, D.; ZAMORA, H.; BLANCO-LABRA, A. 2010. An inhibitor from Lupinus bogotensis seeds effective againsts aspartic proteases from Hypothemus hampei. Phytochemistry 71: 923-929.

NAVARRO, .L; GÓNGORA, C.; BENAVIDES, P. 2010. Single nucleotide polymorphism detection at the Hypothenemus hampei

Rdl gene by allele-specific PCR amplification with T -shift primers. Pesticide Biochemistry and Physiology 97: 204-208.

OLIVEIRA, A. S.; PEREIRA, R. A.; LIMA, L. M.; MORAIS, A. H. A.; MELO, F. R.; FRANCO, O. L.; BLOCH, Jr. C.; GROSSIDE-SÁ, M. F.; SALES, M. P. 2002. Activity toward bruchid pest of a Kunitz-type inhibitor from seeds of the algarroba tree (Prosopis juliflora D. C.). Pesticide Biochemistry and Physiology 72: 122-132.

OSSA, G. A.; BUSTILLO, A. E.; VALENCIA, A. 2000. Determinación del pH en el fluido digestivo de larvas y adultos de Hypothenemus hampei (Coleoptera: Scolytidae). Revista Cenicafé 51: 97-101.

PRASAD, E. R.; DUTTA-GUPTA, A.; PADMASREE, K. 2010. Insecticidal potential of Bowman-Birk proteinasa inhibitors from red gram (Cajanus cajan) and black gram (Vigna mungo) against lepidopteran insect pest. Pesticide Biochemistry and Physiology 98: 80-88.

PRECIADO, D. P.; BUSTILLO, A. E.; VALENCIA, A. 2000. Caracterización parcial de una proteinasa digestiva proveniente de la broca del café (Coleóptera: Scolytidae). Revista Cenicafé 51. 20-27.

RABOSSI, A.; STOKA, V.; PUIZDAR, V.; TURK, V.; QUESADAALLUE, L. A. 2004. Novel aspartyl proteinase associated to fat body histolysis during Ceratitis capitate early metamorphosis. Archives Insect Biochemistry Physiology 57: 51-67.

RODRÍGUEZ, M. L.; ALVES, V.; GRACAS, M.; RICHARDSON, M. 2007. Characterization of a Kunitz trypsin inhibitor with a single disulfide bridge from seeds of Inga laurina (Sw.) Willd. Phytochemistry 68: 1104-1111.

RYAN, C. A. 1990. Proteinase inhibitors in plants: genes for improving defenses against insects and pathogens. Annual Review of Phytopathology 28. 425-449.

SALES, M. P.; ANDRADE, L. B. S.; ARY, M. B.; MIRANDA, M. R. A.; TEXEIRA, F. M.; OLIVEIRA, A. S.; FERNANDES, K. V. S; XAVIER-FILHO, J. 2005. Performance of bean bruchids (Coleoptera: Bruchidae) reared on resistant (IT81D-1045) and susceptible (Epace 10) Vigna unguiculata seeds: Relationship with trypsin inhibitor and vicilin excretion. Comparative Biochemistry and Physiology. Part A: Molecular & Integrative Physiology 142: 422-426.

SCARAFONI, A.; CONSONNI, A.; GALBUSERA, V.; NEGRI, A.; TEDESCHI, G.; RASMUSSEN, P.; MAGNI, C.; DURANTI, M. 2008. Identification and characterization of a Bowman-Birk inhibitor active towards trypsin but not chimotrypsin in Lupinus albus seeds. Phytochemistry 69: 1820-1825.

SCHÄGGER, H.; VON JAGOW, G. 1987. Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for separation of protein in the range of 1 to 100 kDa. Analytical Biochemistry 166: 368-379.

SCHWERTZ, G. W.; TAKENAKA, Y.; 1955. A spectrophotometric determination of trypsin and chymotripsin activity. Biochimica et Biophysica Acta 16: 571-575.

SILVA, C. P.; XAVIER-FILHO, J. 1991. Comparison between the levels of aspartic and cysteine proteinases of the larval midguts of Callosobruchus maculatus (F.) and Zabrotes subfasciatus (Boh.) (Coleoptera: Bruchidae). Comparative Biochemistry and Physiology. Part B: Biochemistry & Molecular Biology 99: 529-533.

SUMIKAWA, J. T.; DE BRITO, M. V.; MACEDO M. L.; UCHOA, A. F.; MIRANDA, A.; ARAUJO, A. P.; SILVA-LUCCA, R. A.; SAMPAIO, M. U.; OLIVA, M. L. 2010. The defensive functions of plant inhibitors are not restricted to insect enzyme inhibition. Phytochemistry 71: 214-220.

TAMHANE,V. A.; GIRI, A. P.; SAINANI, M. N.; GUPTA, V. S. 2007. Diverse forms of Pin-II family proteinase inhibitors from Capsicum annum adversely affect the growth and development of Helicoverpa armigera. Gene 403: 29-38.

TELANG, M.; SRINIVASAN,A.; PATANKAR,A.; HARSULKAR, A.; JOSHI, V.; DAMLE, A.; DESHPANDE, V.; SAINANI, M.; RANJEKAR, P. K.; GUPTA, G.; BIRAH, A.; RANI, S.; KACHOLE, M.; GIRI, A. P.; GUPTA, V.; BIRAH, A.; RANI, S.; KACHOLE, M.; GIRI, A. P.; GUPTA, V. 2003. Bitter gourd proteinase inhibitors: potential growth inhibitors of Helicoverpa armigera and Spodoptera litura. Phytochemistry 63. 643-652.

VALDES, S.; SEGURA, M.; CHAGOLLA, A.; VERVER, A.; MARTÍNEZ, N.; BLANCO, A. 1993. Purification, Characterization, and complete amino sequence of a trypsin inhibitor from Amaranth (Amaranthus hypochondriacus) seeds. Plant Physiology 103: 1407-1412.

VALENCIA, A.; BUSTILLO, A.; OSSA, G.; CHRISPEELS, M. 2000. a-Amylases of the coffee berry borer (Hypothenemus hamperi) and their inhibition by two plant amylase inhibitors. Insect Biochemistry and Molecular Biology 30: 207-213.

VALENCIA, A.; ARBOLEDA, J. W. 2005. Digestion of the inhibitor aAI by Hypothenemus hampei aspartic proteinases. Revista Colombiana de Entomología 31: 117-121.

VALENCIA, A.; ARBOLEDA, J. W.; GROSSI-DE-SÁ. 2007. Detection of a-amylase inhibitors by a zymography method, performed in isoelectric focusing electrophoretic phastGels. Analytical Biochemistry 366: 113-115.

VÁSQUEZ, M.; SMITH, R. H.; MARTÍNEZ, N. A.; BLANCO, A. 1999. Enzymatic differences in the digestive system of the adult and larva of Prostephanus truncates (Horn) (Coleoptera: Bostrichidae). Journal of Stored Products Research 35: 167-174.

VILLALBA, D. A.; BUSTILLO, A. E.; CHÁVES, B. 1995. Evaluación de insecticidas para el control de la broca del café en Colombia. Revista Cenicafé 46: 152-163.

WILHITE, S. E.; ELDEN, T. C.; BRZIN, J.; SMIGOCKI, A. C. 2000. Inhibition of cysteine and aspartyl proteinases in the alfalfa weevil midgut with biochemical and plan-derived proteinase inhibitors. Insect Biochemistry and Molecular Biology 30: 1181-1188.

WOLFSON, J. L.; MURDOCK, L. L. 1987. Supression of larval Colorado beetle growth and development by digestive proteinase inhibitors. Entomologia Experimentalis et Applicata 44: 235-240.

YUNES, A. N. A.; ANDRADE, M. T.; SALES, M. P.; MORAIS, R. A.; FERNANDES, K. V. S.; GOMES, V. M.; XAVIER-FILHO, J. 1998. Legume seed vicilins (7S storage proteins) interfere with the development of the cowpea weevil [Callosobruchus maculatus (F)]. Journal of the Science of Food and Agriculture 76: 111-116.

ZOR, T.; SELINGER, Z. 1996. Linearization of the Bradford protein assay increases its sensitivity: Theoretical and experimental studies. Analytical Biochemistry 236: 302-308.

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 Biochemistry and Molecular Biology 12: 135-145.