A three carbon source feeding strategy for hyaluronic acid production in recombinant escherichia coli
Armento, Donaldson Joseph
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The primary goal of this research was to improve hyaluronic acid production in recombinant E. coli. Hyaluronic acid (HA) is a biopolymer found throughout the human body that has a number of medical applications. A codon optimized hasA gene from Streptococcus zooepidemicus was cloned into wild-type and mutant E. coli MG 1655 strains to allow for HA production. The mutant strains lacked pgi (MEC367), nagB (MEC700), or both genes (MEC526). These genes were selectively knocked out in order to minimize metabolic competition to hyaluronic acid production. The strains were then screened for HA production in shake flasks using 50 mL defined media containing single carbon sources: glucose, fructose, and n-acetylglucosamine, as well as combinations of two or three carbon sources. Four carbon source/strain combinations were then selected to be used in 1-L batch fermentations. MEC 526 grown on 95 mM glucose and 5 mM NAG achieved the highest HA titer of 35.8 mg/L from the 1-L batch fermentation. Utilizing multiple carbon sources improved HA production in all strains. Additionally, MEC 526 which contained both knockouts had higher HA production than the wild-type strain in all experiments.