Production of β-carotene and Acetate in Recombinant Escherichia coli with or without Mevalonate Pathway at Different Culture Temperature or pH

Abstract

Natural beta-carotene has received much attention as consumers have become more health conscious. Its production by various microorganisms including metabolically engineered Escherichia coli or Saccharomyces cerevisiae has been attempted. We successfully created a recombinant E. coli with an engineered whole mevalonate pathway in addition to beta-carotene biosynthetic genes and evaluated the engineered cells from the aspects of metabolic balance between central metabolism and beta-carotene production by comparison with conventional beta-carotene producing recombinant E. coli (control) utilizing a native methylerythritol phosphate (MEP) pathway using bioreactor cultures generated at different temperatures or pHs. Better production of beta-carotene was obtained in E. coli cultured at 37 degrees C than at 25 degrees C. A two-fold higher titer and 2.9-fold higher volumetric productivity were obtained in engineered cells compared with control cells. Notably, a marginal amount of acetate was produced in actively growing engineered cells, whereas more than 8 g/L of acetate was produced in control cells with reduced cell growth at 37 degrees C. The data indicated that the artificial operon of the whole mevalonate pathway operated efficiently in redirecting acetyl-CoA into isopentenyl pyrophosphate (IPP), thereby improving production of beta-carotene, whereas the native MEP pathway did not convert a sufficient amount of pyruvate into IPP due to endogenous feedback regulation. Engineered cells also produced lycopene with a reduced amount of beta-carotene in weak alkaline cultures, consistent with the inhibition of lycopene cyclase.