Inducible expression of a toxic poliovirus membrane protein in Escherichia coli: comparative studies using different expression systems based on T7 promoters.
The poliovirus 3AB gene has been cloned and overproduced in T7 expression vectors using different approaches to allow reduction of basal levels of expression. Expression of the poliovirus 3AB gene is highly toxic for E. coli cells, due to drastic changes induced in membrane permeability of the bacteria that lead to cell lysis when the T7 lysozyme is present. The best production of 3AB was achieved with the T7/lac system in cells lacking T7 lysozyme, where this toxic protein was synthesized to high levels and during several hours in the absence of cell lysis. These results show the efficient synthesis of a highly damaging membrane protein and open the possibility to apply heterologous gene expression in E. coli to other lytic proteins.
Lama
J
J
Centro de Biología Molecular, Universidad Autónoma de Madrid, Canto Blanco.
Carrasco
L
L
D017123:Bacteriophage T7 / Q000235:genetics; D001483:Base Sequence; D002463:Cell Membrane Permeability / Q000187:drug effects; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005822:Genetic Vectors / Q000235:genetics; D007763:Lac Operon / Q000235:genetics; D008969:Molecular Sequence Data; D009113:Muramidase / Q000235:genetics; D017955:Poliovirus / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D012097:Repressor Proteins / Q000235:genetics; D014763:Viral Matrix Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology
D011994:Recombinant Proteins; D012097:Repressor Proteins; D014763:Viral Matrix Proteins; D009113:Muramidase