Expression of human growth hormone in silkworm larvae through recombinant Bombyx mori nuclear polyhedrosis virus.
Expression of human growth hormone in silkworm larvae through recombinant Bombyx mori nuclear polyhedrosis virus.
复制标题
通过重组家蚕核多角体病毒在家蚕幼虫中表达人生长激素。
DOI：
10.1006/prep.1996.0037
复制
发表时间：
1996
影响因子：
1.6
通讯作者：
K. Gopinathan
中科院分区：
文献类型：
--
作者：
S. Sumathy;V. Palhan;K. Gopinathan
我们已经产生了重组家蚕核型多角体病毒，vBmhGH，窝藏全长人生长激素基因（2.4 kb的基因组DNA，4个内含子和信号肽序列）的多角体蛋白启动子的控制下。用重组病毒感染培养的BmN细胞显示存在对应于真实生长激素mRNA及其不完全加工的前体的RNA。重组病毒感染的BmN细胞的蛋白质的电泳分析和免疫沉淀揭示了生长激素蛋白的存在。家蚕幼虫感染vBmhGH后，其蛋白质合成并分泌到血淋巴中。重组人生长激素在放射受体竞争结合试验中具有生物活性。通过一步免疫亲和层析将分泌的蛋白分离并纯化至均一，至比活性为2.4 × 10（4）U/mg。重组hGH保留了天然肽的免疫学和生物学特性。我们的结论是，BmNPV载体可以成功地用于表达含有多个内含子的染色体基因。
We have generated a recombinant Bombyx mori nuclear polyhedrosis virus, vBmhGH, harboring the full length human growth hormone gene (2.4-kb genomic DNA, with four introns and the signal peptide sequences) under the control of the polyhedrin promoter. BmN cells in culture infected with the recombinant virus showed the presence of RNA corresponding to the authentic growth hormone mRNA as well as its incompletely processed precusor. Electrophoretic analysis and immunoprecipitation of proteins of recombinant virus-infected BmN cells revealed the presence of the growth hormone protein. Infection of silkworm larvae with vBmhGH led to the synthesis and efficient secretion of the protein into hemolymph. The recombinant human growth hormone was biologically active in a radioreceptor competition binding assay. The secreted protein was isolated and purified to homogeneity by a single step immunoaffinity chromatography, to a specific activity of 2.4 x 10(4) U/mg. The recombinant hGH retained the immunological and biological properties of the native peptide. We conclude that BmNPV vectors can be used successfully for expressing chromosomal genes harboring multiple introns.