Breugelmans Bert, Simonet Gert, van Hoef Vincent, Van Soest Sofie, Smagghe Guy, Vanden Broeck Jozef
Department of Animal Physiology and Neurobiology, Zoological Institute K.U. Leuven, Naamsestraat 59, B-3000 Leuven, Belgium.
Insect Biochem Mol Biol. 2009 Jul;39(7):430-9. doi: 10.1016/j.ibmb.2009.03.005. Epub 2009 Apr 11.
Members of the pacifastin family have been characterized as serine peptidase inhibitors (PI), but their target enzyme(s) are unknown in insects. So far, the structural and biochemical characteristics of pacifastin-like PI have only been studied in locusts. Here we report the molecular identification and functional characterization of a pacifastin-like precursor in a lepidopteran insect, i.e. the silkworm Bombyx mori. The bmpp-1 gene contains 17 exons and codes for two pacifastin-related precursors of different length. The longest splice variant encodes 13 inhibitor domains, more than any other pacifastin-like precursor in arthropods. The second transcript lacks two exons and codes for 11 inhibitor domains. By studying the expression profile of the Bombyx pacifastin-like gene a different expression pattern for the two variants was observed suggesting functional diversification. Next, several PI domains of BMPP-1 were produced and, contrary to locust pacifastin peptides, they were found to be potent inhibitors of both bovine trypsin and chymotrypsin. Surprisingly, the same Bombyx PI are only weak inhibitors of endogenous digestive peptidases, indicating that other peptidases are the in vivo targets. Interestingly, the Bombyx PI inhibit a fungal trypsin-like cuticle degrading enzyme, suggesting a protective function for BMPP-1 against entomopathogenic fungi.
pacifastin家族成员已被鉴定为丝氨酸肽酶抑制剂（PI），但其在昆虫中的靶标酶尚不清楚。到目前为止，类pacifastin PI的结构和生化特性仅在蝗虫中进行了研究。在此，我们报告了一种鳞翅目昆虫（即家蚕Bombyx mori）中类pacifastin前体的分子鉴定和功能表征。bmpp - 1基因包含17个外显子，编码两种不同长度的与pacifastin相关的前体。最长的剪接变体编码13个抑制剂结构域，比节肢动物中任何其他类pacifastin前体都多。第二个转录本缺少两个外显子，编码11个抑制剂结构域。通过研究家蚕类pacifastin基因的表达谱，观察到这两种变体具有不同的表达模式，表明存在功能分化。接下来，制备了BMPP - 1的几个PI结构域，与蝗虫pacifastin肽不同，发现它们是牛胰蛋白酶和胰凝乳蛋白酶的有效抑制剂。令人惊讶的是，相同的家蚕PI对内源消化肽酶只是弱抑制剂，这表明其他肽酶是体内靶标。有趣的是，家蚕PI抑制一种真菌类胰蛋白酶角质层降解酶，表明BMPP - 1对昆虫病原真菌具有保护作用。
