Crystal structure of Bombyx mori arylphorins reveals a 3:3 heterohexamer with multiple papain cleavage sites

In holometabolous insects, the accumulation and utilization of storage proteins (SPs), including arylphorins and methionine-rich proteins, are critical for the insect metamorphosis. SPs function as amino acids reserves, which are synthesized in fat body, secreted into the larval hemolymph and taken up by fat body shortly before pupation. However, the detailed molecular mechanisms of digestion and utilization of SPs during development are largely unknown. Here, we report the crystal structure of Bombyx mori arylphorins at 2.8 angstrom, which displays a heterohexameric structural arrangement formed by trimerization of dimers comprising two structural similar arylphorins. Our limited proteolysis assay and microarray data strongly suggest that papain-like proteases are the major players for B. mori arylphorins digestion in vitro and in vivo. Consistent with the biochemical data, dozens of papain cleavage sites are mapped on the surface of the heterohexameric structure of B. mori arylphorins. Hence, our results provide the insightful information to understand the metamorphosis of holometabolous insects at molecular level.

在全变态昆虫中，包括芳基脂蛋白和富含甲硫氨酸的蛋白质在内的储存蛋白（SPs）的积累和利用对昆虫的变态至关重要。储存蛋白作为氨基酸储备，在脂肪体中合成，分泌到幼虫血淋巴中，并在化蛹前不久被脂肪体吸收。然而，在发育过程中储存蛋白消化和利用的详细分子机制在很大程度上是未知的。在此，我们报道了家蚕芳基脂蛋白在2.8埃的晶体结构，其呈现出一种由两个结构相似的芳基脂蛋白组成的二聚体三聚化形成的异六聚体结构排列。我们的有限蛋白水解实验和微阵列数据有力地表明，类木瓜蛋白酶是家蚕芳基脂蛋白在体外和体内消化的主要参与者。与生化数据一致，在家蚕芳基脂蛋白的异六聚体结构表面定位到了数十个类木瓜蛋白酶切割位点。因此，我们的研究结果为在分子水平上理解全变态昆虫的变态提供了有深刻见解的信息。