Expression of human growth hormone in silkworm larvae through recombinant Bombyx mori nuclear polyhedrosis virus.
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.
Sumathy
S
S
Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India.
Palhan
V B
VB
Gopinathan
K P
KP
D000818:Animals; D012831:Bombyx / Q000235:genetics*; D002460:Cell Line; D005260:Female; D015870:Gene Expression; D017344:Genes, Insect / Q000235:genetics; D005822:Genetic Vectors; D013006:Growth Hormone / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D066298:In Vitro Techniques; D007814:Larva / Q000235:genetics; D008099:Liver / Q000378:metabolism; D017924:Nucleopolyhedroviruses / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011247:Pregnancy; D051381:Rats; D011986:Receptors, Somatotropin / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics; D014158:Transcription, Genetic
D011986:Receptors, Somatotropin; D011994:Recombinant Proteins; D013006:Growth Hormone