Acetate formation during recombinant protein production in K-12 with an elevated NAD(H) pool.

Acetate formation is a disadvantage in the use of for recombinant protein production, and many studies have focused on optimizing fermentation processes or altering metabolism to eliminate acetate accumulation. In this study, MEC697 (MG1655 ) maintained a larger pool of NAD(H) compared to the wild-type control, and also accumulated lower concentrations of acetate when grown in batch culture on glucose. In steady-state cultures, the elevated total NAD(H) found in MEC697 delayed the threshold dilution rate for acetate formation to a growth rate of 0.27 h. Batch and fed-batch processes using MEC697 were examined for the production of β-galactosidase as a model recombinant protein. Fed-batch culture of MEC697/pTrc99A- compared to MG1655/pTrc99A- at a growth rate of 0.22 h showed only a modest increase of protein formation. However, 1 L batch growth of MEC697/pTrc99A- resulted in 50% lower acetate formation compared to MG1655/pTrc99A- and a two-fold increase in recombinant protein production.