Strain Engineering for High-Level 5-Aminolevulinic Acid Production in Escherichia Coli Publisher Pubmed



Miscevic D¹ ; Mao JY² ; Kefale T^{1, 3} ; Abedi D^{1, 4} ; Mooyoung M¹ ; Perry Chou C¹
Source: Biotechnology and Bioengineering Published:2021

Abstract

Herein, we report the development of a microbial bioprocess for high-level production of 5-aminolevulinic acid (5-ALA), a valuable non-proteinogenic amino acid with multiple applications in medical, agricultural, and food industries, using Escherichia coli as a cell factory. We first implemented the Shemin (i.e., C4) pathway for heterologous 5-ALA biosynthesis in E. coli. To reduce, but not to abolish, the carbon flux toward essential tetrapyrrole/porphyrin biosynthesis, we applied clustered regularly interspersed short palindromic repeats interference (CRISPRi) to repress hemB expression, leading to extracellular 5-ALA accumulation. We then applied metabolic engineering strategies to direct more dissimilated carbon flux toward the key precursor of succinyl-CoA for enhanced 5-ALA biosynthesis. Using these engineered E. coli strains for bioreactor cultivation, we successfully demonstrated high-level 5-ALA biosynthesis from glycerol (~30 g L−1) under both microaerobic and aerobic conditions, achieving up to 5.95 g L−1 (36.9% of the theoretical maximum yield) and 6.93 g L−1 (50.9% of the theoretical maximum yield) 5-ALA, respectively. This study represents one of the most effective bio-based production of 5-ALA from a structurally unrelated carbon to date, highlighting the importance of integrated strain engineering and bioprocessing strategies to enhance bio-based production. © 2020 Wiley Periodicals LLC