Development of a high cell-density fed-batch bioprocess for the heterologous production of 6-deoxyerythronolide B in Escherichia coli.
A robust high cell-density fed-batch bioprocess was developed for the heterologous production of 6-deoxyerythronolide B (6-dEB), the macrocyclic core of the antibiotic erythromycin, with a recombinant Escherichia coli strain expressing the 6-deoxyerythronolide B synthase (DEBS) from Saccharopolyspora erythraea. Initial evaluation of the E. coli strain in a 5-l bioreactor with the addition of exogenous propionate for polyketide biosynthesis resulted in a maximum cell density of 30 g l(-1) (OD600 approximately 60) and the production of 700 mg l(-1) of 6-dEB. Retention of the two plasmids harboring the heterologous genes was maintained between 90 and 100% even in the absence of antibiotic selection. However, the accumulation of excess ammonia in the culture medium was found to significantly decrease the productivity of the cells. Through optimization of the medium composition and fermentation conditions, the maximum cell density was increased by two-fold, and a final titer of 1.1 g l(-1) of 6-dEB was achieved. This represents an 11-fold improvement compared to the highest reported titer of 100 mg l(-1) with E. coli as the production host.
Lau
Janice
J
Department of Process Science, Kosan Biosciences, Inc., 3832 Bay Center Place, Hayward, CA 94545, USA. lau@kosan.com
Tran
Carnie
C
Licari
Peter
P
Galazzo
Jorge
J
D000641:Ammonia / Q000494:pharmacology; D019149:Bioreactors; D001709:Biotechnology / Q000379:methods*; D002452:Cell Count; D003470:Culture Media / Q000737:chemistry*; D004917:Erythromycin / Q000031:analogs & derivatives* / Q000096:biosynthesis*; D004926:Escherichia coli / Q000378:metabolism*; D005285:Fermentation; D011422:Propionates / Q000494:pharmacology; D048372:Streptomyces coelicolor / Q000378:metabolism; D013997:Time Factors
D003470:Culture Media; D011422:Propionates; C034277:6-deoxyerythronolide B; D004917:Erythromycin; D000641:Ammonia; C029658:propionic acid