Overcoming inclusion body formation in a high-level expression system.
Attempts at overexpressing T4-phage deoxycytidylate deaminase using the pET3c/BL21(DE3)/pLysS system resulted in this enzyme being part of an inactive inclusion-body complex. However, by employing an enriched growth medium it was found that the deaminase could be induced in a soluble active form to at least 20% of this organism's cellular protein. Insoluble inclusion bodies were obtained with less rich media. This procedure was employed successfully with other highly expressed proteins that formed inclusion bodies. The use of a rich growth medium during the course of protein induction may be a valuable adjunct to limiting inclusion body formation with this as well as other expression systems.
Moore
J T
JT
Wadsworth Center for Laboratories and Research, New York State Department of Health, Albany.
Uppal
A
A
Maley
F
F
Maley
G F
GF
D017122:Bacteriophage T4 / Q000201:enzymology* / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D003470:Culture Media; D003844:DCMP Deaminase / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D005814:Genes, Viral / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D007181:Inclusion Bodies, Viral / Q000201:enzymology*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis; D015678:Viral Structural Proteins / Q000235:genetics
D003470:Culture Media; D011994:Recombinant Proteins; D015678:Viral Structural Proteins; D003844:DCMP Deaminase