Molecular mechanisms of biosynthesis and degradation of yolk proteins of the silkworm,Bombyx mori

家蚕的卵黄蛋白由三种主要蛋白组成，其中卵特异性蛋白(ESP)的行为与其他蛋白不同。本研究旨在阐明该蛋白生物合成和降解的分子机制。从基因组文库中分离出ESP基因，并对其部分结构进行了鉴定。构建了ESP的cDNA，并进行了测序。从核苷酸序列推导出完整的氨基酸序列。体外翻译实验表明，ESP被翻译为一个新生的节省多肽的序列。翻译后修饰通过信号切割、糖基化、磷酸化依次发生。ESP含有相当数量的锌，这表明ESP与核酸相互作用。在胚胎发育过程中，ESP被一种独特的蛋白酶依次降解为三种多肽。该酶对ESP具有较高的特异性，与其他蛋黄蛋白无明显反应。通过分析赖氨酸^&lt；114&gt；和精氨酸^&lt；210&gt；的N末端氨基酸序列，确定了裂解位点。该酶的活性随胚胎发生的不同而呈暂时性变化。这种活动的发育曲线与mRNA活性的变化很好地对应。体外翻译实验表明，该酶蛋白是以新生多肽的形式合成，然后通过信号切割加工成成熟的蛋白质。新生肽具有与加工蛋白相当的生物活性。这是首次报道具有生物活性的翻译产物。

Yolk proteins the silkworm, Bombyx mori consist of three major proteins and among them, egg-specific (ESP) behaves differently from others. The present study was carried out to elucidate the molecular mechanisms of biosynthesis and degradation of this protein. From the genomic library, ESP gene was isolated and the partial structure was carified. cDNA of ESP was constracted and sequenced. The complete amino acid sequence was deduced from the nucleotids sequence. The in vitro translation experiment concluded that esp was translated as a nascent peptide-aving segnal sequence. The post-translational modification occurred sequentially through signal cleavage, glycosylation, phosphorylation. ESP contains appreciable amounts of zinc, which suggest that ESP interacts with nucleic acids.During embryogenesis, ESP is degraded sequentially into three peptides by a unique protease. This protease had the high specificity to ESP without atacking to the other yolk proteins. Cleavage sites were identified by analysing N-termina amino acid sequence as at the carboxyl site of lysine^<114> and argine^<210>. The activity of this protease chenged temporaly according to the embryogenesis. This developmental profile in activity well corresponded to the changes in mRNA activity. The in vitro translation experiment showed that the enzyme protein is synthesized as a nascent peptide and then processed into a mature form of protein through signal cleavage. The nascent peptide showed the biological activity which was comparable to the processed proteins. This is the first report of the biologically active products of translation.