Zibaee Arash, Bandani Ali Reza, Fazeli-Dinan Mahmoud, Zibaee Idin, Sendi Jalal Jalali, Maleki Feizollah A
Department of Plant Protection, College of Agriculture, University of Guilan, Rasht, Iran.
Arch Insect Biochem Physiol. 2011 Sep;78(1):1-16. doi: 10.1002/arch.20435. Epub 2011 Jun 15.
The rice green semi looper, Naranga aenescens Moore (Lepidoptera: Noctuidae) causes severe damage to rice fields in Eastern Asia and Middle East. We demonstrate that two types of serine proteases are active in the midgut of the third instar larvae of N. aenescens, but trypsin-like proteases are considerably more active than chymotrypsin-like proteases. To develop better control strategies, purification and biochemical characterization of a major trypsin-like digestive protease from the midgut of the third instar larvae of N. aenescens was achieved by gel filtration and anion exchange chromatography. After the final purification step, the enzyme was purified 9.62-fold with a recovery of 16.1% and a specific activity of 4.12 U/mg protein and a molecular mass of approximately 88.5 kDa. Biochemical characterization indicated that the purified protease had highest activity at pH 10 and 30°C and was stable for up to 6 h under those conditions. Divalent cations, especially Ca2+, Mg2+, and Cu2+, increased the enzyme activity and synthetic inhibitors that target trypsin-like activity caused a significant reduction in caseinolytic activity. These data may be used to develop inhibitors that decrease the damage of N. aenescens to rice cultivars in the field.
稻绿夜蛾（Naranga aenescens Moore，鳞翅目：夜蛾科）对东亚和中东的稻田造成严重破坏。我们证明，两种类型的丝氨酸蛋白酶在稻绿夜蛾三龄幼虫的中肠中具有活性，但类胰蛋白酶比类糜蛋白酶的活性要高得多。为了制定更好的防治策略，通过凝胶过滤和阴离子交换色谱法，对稻绿夜蛾三龄幼虫中肠中的一种主要类胰蛋白酶消化蛋白酶进行了纯化和生化特性分析。在最后一步纯化后，该酶的纯化倍数为9.62倍，回收率为16.1%，比活性为4.12 U/mg蛋白，分子量约为88.5 kDa。生化特性分析表明，纯化后的蛋白酶在pH 10和30°C时活性最高，在这些条件下可稳定长达6小时。二价阳离子，尤其是Ca2+、Mg2+和Cu2+，可提高该酶的活性，而针对类胰蛋白酶活性的合成抑制剂会导致酪蛋白水解活性显著降低。这些数据可用于开发抑制剂，以减少稻绿夜蛾对田间水稻品种的损害。
