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首页> 外文期刊>Comparative and functional genomics >Expression, Purification, and Characterization of Ras Protein (BmRas1) fromBombyx mori
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Expression, Purification, and Characterization of Ras Protein (BmRas1) fromBombyx mori

机译：家蚕Ras蛋白（BmRas1）的表达，纯化和鉴定

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摘要

The Ras subfamily is the member of small G proteins superfamily involved in cellular signal transduction. Activation of Ras signaling causes cell growth, differentiation, and survival.Bombyx moriRas-like protein (BmRas1) may belong to the Ras subfamily. It contained an H-N-K-Ras-like domain. The BmRas1 mRNA consisted of 1459 bp. The open reading frame contained 579 bp, encoding 192 amino acids. The protein had such secondary structures asα-helices, extended strand, and random coil. BmRas1 was expressed successfully inE. coliBL21. The recombinant protein was purified with metal-chelating affinity chromatography. The GTPase activity of purified protein was determined by FeSO4-(NH4)2MoO4assay. The results showed that purified recombinant protein had intrinsic activity of GTPase. High titer polyclonal antibodies were generated by New Zealand rabbit immunized with purified protein. The gene expression features of BmRas1 at different stages and in different organs of the fifth instar larvae were analyzed by Western blot. The results showed that BmRas1 was expressed highly in three development stages including egg, pupae, and adult, but low expression in larva. BmRas1 was expressed in these tissues including head, malpighian tubule, genital gland, and silk gland. The purified recombinant protein would be utilized to further function studies of BmRas1.
机译：Ras家族是参与细胞信号转导的小G蛋白超家族的成员。 Ras信号的激活导致细胞的生长，分化和存活。BombyxmoriRas样蛋白（BmRas1）可能属于Ras家族。它包含一个H-N-K-Ras样结构域。 BmRas1 mRNA由1459bp组成。开放阅读框包含579 bp，编码192个氨基酸。该蛋白质具有二级结构，例如α-螺旋，延伸链和无规卷曲。 BmRas1在E中成功表达。大肠杆菌BL21。用金属螯合亲和色谱法纯化重组蛋白。通过FeSO4-（NH4）2MoO4测定法测定纯化蛋白的GTPase活性。结果表明，纯化的重组蛋白具有GTPase的固有活性。用纯化蛋白免疫的新西兰兔产生高滴度的多克隆抗体。用Western blot分析BmRas1基因在五龄幼虫不同阶段，不同器官的基因表达特征。结果表明，BmRas1在卵，p和成虫三个发育阶段均高表达，而在幼虫中低表达。 BmRas1在这些组织中表达，包括头部，马尔福氏小管，生殖腺和丝腺。纯化的重组蛋白将用于BmRas1的进一步功能研究。

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