A new combination approach to extracellular production of 5-aminolevulinic acid for purification and application in alleviating cadmium-induced oxidative stress in maize

Abstract

5-Aminolevulinic acid (ALA) is widely applied in agriculture, animal husbandry, medicine, and often manufactured in Escherichia coli for overexpressing ALA synthase (ALAS) from α-proteobacteria. For enhancing extracellular ALA production, several approaches have been exploited. Here, we developed and identified a new combination strategy to increase ALA production in E. coli, including selection of the negatively-charged peptide tag as a C-terminal fusion partner for increasing soluble production of the ALAS codon variant from Rhodobacter sphaeroides, mutation of certain residues to increase the ALAS variant activity, optimization of the signal sequences to facilitate ALA secretion, down-regulation of the hemB to inhibit ALA transformation in one plasmid expression system, and supply of 4 mM dithiothreitol to the culture to increase cells tolerant to the oxidative stress. Under the specified cultural conditions, ALA yield was up to 3.2 g/L in flash flasks. Compared with the added cadmium-induced stress, simultaneous supply of purified ALA improved maize seedlings growth, decreased contents of malondialdehyde and hydrogen peroxide, and increased peroxidase activity, contents of chlorophylls and proline.