Expression of post-translational processing of preprocecropin A using a baculovirus vector.
A cDNA fragment encoding preprocecropin A was inserted into the baculovirus Autographa californica nuclear polyhedrosis virus downstream of the polyhedrin promoter. The gene was expressed in recombinant-infected last instar larvae of Trichoplusia ni and in diapausing pupae of Hyalophora cecropia. The identity of the recombinant product was established by electrophoresis with detection of antibacterial activity and mass spectrometry. The prepropeptide had been correctly processed including removal of signal peptide and pro-part. Biologically active and amidated cecropin A was exported to the hemolymph. The yield of recombinant protein in H. cecropia reached a level of 600 micrograms/ml hemolymph and about 70% of the material was amidated.
Hellers
M
M
Department of Microbiology, University of Stockholm, Sweden.
Gunne
H
H
Steiner
H
H
D000818:Animals; D023181:Antimicrobial Cationic Peptides; D016367:Baculoviridae / Q000235:genetics*; D015152:Blotting, Northern; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D015870:Gene Expression; D005822:Genetic Vectors; D007301:Insect Hormones / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D019476:Insect Proteins; D007313:Insecta; D013058:Mass Spectrometry; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics*; D011499:Protein Processing, Post-Translational; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D014162:Transfection
D023181:Antimicrobial Cationic Peptides; C059037:CecA2 protein, Bombyx mori; D007301:Insect Hormones; D019476:Insect Proteins; D011498:Protein Precursors; D011994:Recombinant Proteins; D012313:RNA; C031161:cecropin A