Franco Octávio L, dos Santos Roseane C, Batista João A N, Mendes Ana Cristina M, de Araújo Marcus Aurélio M, Monnerat Rose G, Grossi-de-Sá Maria Fátima, de Freitas Sonia M
EMBRAPA Recursos Geneticos e Biotecnologia, 70770 900, Brasi;lia-DF, Brazil.
Phytochemistry. 2003 Jun;63(3):343-9. doi: 10.1016/s0031-9422(03)00108-0.
The cotton boll weevil Anthonomus grandis (Boheman) is one of the major pests of cotton (Gossypium hirsutum L.) in tropical and sub-tropical areas of the New World. This feeds on cotton floral fruits and buds causing severe crop losses. Digestion in the boll weevil is facilitated by high levels of serine proteinases, which are responsible for the almost all proteolytic activity. Aiming to reduce the proteolytic activity, the inhibitory effects of black-eyed pea trypsin/chymotrypsin inhibitor (BTCI), towards trypsin and chymotrypsin from bovine pancreas and from midguts of A. grandis larvae and adult insects were analyzed. BTCI, purified from Vigna unguiculata (L.) seeds, was highly active against different trypsin-like proteinases studied and moderately active against the digestive chymotrypsin of adult insects. Nevertheless, no inhibitory activity was observed against chymotrypsin from A. grandis larval guts. To test the BTCI efficiency in vivo, neonate larvae were reared on artificial diet containing BTCI at 10, 50 and 100 microM. A reduction of larval weight of up to approximately 54% at the highest BTCI concentration was observed. At this concentration, the insect mortality was 65%. This work constitutes the first observation of a Bowman-Birk type inhibitor active in vitro and in vivo toward the cotton boll weevil A. grandis. The results of bioassays strongly suggest that BTCI may have potential as a transgene protein for use in engineered crop plants modified for heightened resistance to the cotton boll weevil.
棉铃象甲Anthonomus grandis（博赫曼）是新大陆热带和亚热带地区棉花（陆地棉L.）的主要害虫之一。它以棉花的花、果实和芽为食，会造成严重的作物损失。棉铃象甲的消化过程由高水平的丝氨酸蛋白酶促进，这些酶几乎负责所有的蛋白水解活性。为了降低蛋白水解活性，分析了黑眼豆胰蛋白酶/胰凝乳蛋白酶抑制剂（BTCI）对牛胰腺以及棉铃象甲幼虫和成虫中肠的胰蛋白酶和胰凝乳蛋白酶的抑制作用。从豇豆（L.）种子中纯化得到的BTCI对所研究的不同胰蛋白酶样蛋白酶具有高活性，对成虫的消化胰凝乳蛋白酶具有中等活性。然而，未观察到对棉铃象甲幼虫中肠胰凝乳蛋白酶的抑制活性。为了测试BTCI在体内的效率，将新生幼虫饲养在含有10、50和100微摩尔BTCI的人工饲料上。在最高BTCI浓度下，观察到幼虫体重减轻高达约54%。在此浓度下，昆虫死亡率为65%。这项工作首次观察到一种Bowman-Birk型抑制剂在体外和体内对棉铃象甲A. grandis具有活性。生物测定结果强烈表明，BTCI可能有潜力作为一种转基因蛋白用于经过基因改造以增强对棉铃象甲抗性的作物。
