A recombinant lipoprotein antigen against Lyme disease expressed in E. coli: fermentor operating strategies for improved yield.
Decorin-binding lipoprotein, lpp-DBP, a bacterial surface adhesin, shows promise as a vaccine against Lyme disease. It is expressed in recombinant E. coli as an undesirable 20.5 KDa apoprotein that is subsequently lipidated in vivo to the desired 22 KDa lpp-DBP form. This study defines fermentation conditions for maximizing lpp-DBP yield. Super broth medium, a low post-induction temperature (30 degrees C), and a glucose feed based on dissolved oxygen resulted in high lpp-DBP yield and minimized apoprotein formation. Since cells lysed within 2-3 h after induction, the cell yield was maximized by growing cells to high cell density prior to induction. Compared to a glucose feed based on maintaining a constant fermentor glucose concentration (Glucose-Stat), feeding based on maintaining a constant dissolved oxygen level (DO-Stat) improved yields. Also, a dissolved oxygen level of 60% (air saturation) was best, as no product degradation was detected by Western blotting and SDS-PAGE. Acetic acid levels under both modes of glucose feed were sufficiently low, and no adverse growth effects were observed.
Madurawe
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RD
Bioprocess Scale-Up Facility, Engineering Research Center, University of Maryland, College Park, Maryland 20742, USA.
Chase
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TE
Tsao
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EI
Bentley
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WE
D000942:Antigens, Bacterial / Q000235:genetics*; D019149:Bioreactors; D002244:Carbon; D003470:Culture Media; D004926:Escherichia coli / Q000235:genetics*; D005285:Fermentation; D005947:Glucose; D008074:Lipoproteins / Q000235:genetics*; D008193:Lyme Disease / Q000276:immunology*; D011994:Recombinant Proteins / Q000235:genetics; D013696:Temperature
D000942:Antigens, Bacterial; D003470:Culture Media; D008074:Lipoproteins; D011994:Recombinant Proteins; D002244:Carbon; D005947:Glucose