Efficient aspartic acid production by a psychrophile-based simple biocatalyst

Abstract

We previously constructed a Psychrophile-based Simple bioCatalyst (PSCat) reaction system, in which psychrophilic metabolic enzymes are inactivated by heat treatment, and used it here to study the conversion of aspartic acid from fumaric acid mediated by the activity of aspartate ammonia-lyase (aspartase). In Escherichia coli, the biosynthesis of aspartic acid competes with that of L-malic acid produced from fumaric acid by fumarase. In this study, E. coli aspartase was expressed in psychrophilic Shewanella livingstonensis Ac10 heat treated at 50 °C for 15 min. The resultant PSCat could convert fumaric acid to aspartic acid without the formation of L-malic acid because of heat inactivation of psychrophilic fumarase activity. Furthermore, alginate-immobilized PSCat produced high yields of aspartic acid and could be re-used nine times. The results of our study suggest that PSCat can be applied in biotechnological production as a new approach to increase the yield of target compounds.

Sweet Dreams (Are Made of This): A Review and Perspectives on Aspartic Acid Production

“…Stereospecific, industrial production of L-aspartic acid currently utilizes a one-step reaction of fumaric acid, in the presence of high concentrations of ammonia, to L-aspartic acid via L-aspartate ammonia-lyase, an enzyme also referred to as "L-aspartase" (Figure 1) [7]. L-aspartase can be purified and immobilized in a gel matrix for continuous production or overexpressed in bacterial cells bound to polyurethane carriers [7].…”

“…Stereospecific, industrial production of L-aspartic acid currently utilizes a one-step reaction of fumaric acid, in the presence of high concentrations of ammonia, to L-aspartic acid via L-aspartate ammonia-lyase, an enzyme also referred to as "L-aspartase" (Figure 1) [7]. L-aspartase can be purified and immobilized in a gel matrix for continuous production or overexpressed in bacterial cells bound to polyurethane carriers [7]. Production via immobilized enzymatic conversion or bacterial fermentation utilizes the same enzyme and substrate; however, enzymatic production is favored for its high product concentration, productivity, minimal byproducts, and the ease of downstream processing [2].…”

“…Substrates used for the production of biochemical primary substrate: fumaric acid [7] cofactor: ammonia [7] enzyme: L-aspartate ammonia-lyase [7] Microorganisms used for fermentation Primary industrial species: E. coli and Cornybacterium glutamacium [9] Exploratory species [1]:…”

“…Side products, byproducts, waste products and associated cost Organisms whose genomes also code fumarase (e.g., C. glutamicum, E. coli) produce malic acid from fumarate as a byproduct in effect utilizing substrate and decreasing aspartic acid yield. Without heat treatments Tajima et al (2015) lost 25% of the fumaric acid substrate to malic acid production which translates to significant yield losses [7].…”

Enzymes of industrial interest

“…Free and immobilized thermostable aspartase of B. subtilis YM-55-1, expressed in E. coli, has been used to prepare 500 mM aspartic acid (66 g/L) in a 24 h fermentation (Tajima et al, 2015). Aspartic acid is used to make the synthetic sweetener aspartame and amino acid analogs of C4 building block chemicals, such as 1,4-butanediol, tetrahydrofuran, and gamma-butyrolactone.…”