Li Si, Fei JingJing, Cheng DanDan, Jin Yongfeng, Zhang Wenping, Zhang Yaozhou, Lv Zhengbing
Institute of Biochemistry, College of Life Sciences, Zhejiang Sci-Tech University, Hangzhou, China.
Zhejiang Provincial Key Laboratory of Silkworm Bioreactor and Biomedicine, Zhejiang Sci-Tech University, Hangzhou, China.
Arch Insect Biochem Physiol. 2016 Feb;91(2):109-23. doi: 10.1002/arch.21312. Epub 2015 Dec 17.
FK506 binding proteins (FKBPs) are intracellular receptors of the immunosuppressant FK506 and play important roles in the correct folding of new proteins and the self-assembly of biological macromolecules. FKBP12 is a member of the FKBP family that is widely expressed and highly conserved in many species. In this study, we identified the complete cDNA sequence encoding the FKBP12 ortholog in Bombyx mori, named Bm-FKBP12B (GenBank accession no. DQ443423). Multiple-sequence alignment among different species revealed a high similarity among FKBP12 paralogs and orthologs. Bioinformatics analysis of the Bm-FKBP12B gene showed that it is located on chromosome 20 and consists of three exons and two introns. We cloned, expressed, and purified the Bm-FKBP12B protein in Escherichia coli and generated a specific polyclonal antibody against Bm-FKBP12B. The real-time quantitative reverse-transcription (qRT) PCR and Western blotting results showed that Bm-FKBP12B was present throughout all of the development stages, but it was abundant in the adult and embryo stages. Bm-FKBP12B expression was higher in the silk gland and gut, suggesting that it might play important roles in regulating gene expression in the silk gland and during silk fiber formation. Bm-FKBP12B protein was distributed in the cytoplasm, nucleus, and nuclear membrane.
FK506结合蛋白（FKBPs）是免疫抑制剂FK506的细胞内受体，在新蛋白质的正确折叠和生物大分子的自组装中发挥重要作用。FKBP12是FKBP家族的成员，在许多物种中广泛表达且高度保守。在本研究中，我们鉴定了家蚕中编码FKBP12直系同源物的完整cDNA序列，命名为Bm-FKBP12B（GenBank登录号：DQ443423）。不同物种间的多序列比对显示FKBP12旁系同源物和直系同源物之间具有高度相似性。对Bm-FKBP12B基因的生物信息学分析表明，它位于20号染色体上，由三个外显子和两个内含子组成。我们在大肠杆菌中克隆、表达并纯化了Bm-FKBP12B蛋白，并制备了针对Bm-FKBP12B的特异性多克隆抗体。实时定量逆转录（qRT）PCR和蛋白质印迹结果表明，Bm-FKBP12B在所有发育阶段均有表达，但在成虫和胚胎阶段含量丰富。Bm-FKBP12B在丝腺和肠道中的表达较高，表明它可能在调节丝腺基因表达和丝纤维形成过程中发挥重要作用。Bm-FKBP12B蛋白分布于细胞质、细胞核和核膜中。
