Flow cytometric analysis of metabolic stress effects due to recombinant plasmids and proteins in Escherichia coli production strains.
Overexpression of recombinant proteins in Escherichia coli often leads to a severe growth retardation of the host cells. Using flow cytometry, we analyzed the temporal development of the cellular content of DNA, total protein, and the recombinant product (human superoxide dismutase) in different strains. In cells carrying plasmids utilizing the phage T7 promoter 10 (pET vectors), induction with IPTG leads to an increase in protein content and size, an increase and a wide spreading of DNA content distribution, and a termination of cell division. These effects occurred with pET plasmids with or without an insert, but not with another plasmid which utilizes the tac promoter.
Soriano
E
E
Institut für Angewandte Mikrobiologie, Universität für Bodenkultur, Vienna, Austria.
Borth
N
N
Katinger
H
H
Mattanovich
D
D
D017123:Bacteriophage T7 / Q000235:genetics; D001709:Biotechnology; D004274:DNA, Recombinant / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D013482:Superoxide Dismutase / Q000096:biosynthesis / Q000235:genetics
D004274:DNA, Recombinant; D011994:Recombinant Proteins; D013482:Superoxide Dismutase