General Description

L-Alanine, a crucial compound with widespread applications in various industries, is utilized in clinical medicine, food production, and the development of innovative materials like MGDA and biodegradable polymers. Through metabolic engineering of microorganisms, significant advancements have been made in enhancing L-alanine production efficiency. Strategies such as overexpressing key genes like B. sphaericus L-ALD alaD and minimizing coproduct formation have led to improved yields and purity levels. Genetic engineering efforts in organisms like E. coli have shown promise in achieving high levels of L-alanine production. The implementation of novel genetic switches has allowed for precise control over production, even under challenging conditions. Overall, these advancements highlight the increased potential for sustainable and efficient L-alanine production in the biotechnology industry through innovative metabolic engineering techniques and strain optimization.

Figure 1. L-Alanine

Overview

L-Alanine, a small chiral compound, plays a crucial role in various industries including food, pharmaceuticals, and veterinary fields. Its versatility and widespread use stem from its numerous applications. In the realm of clinical medicine, L-Alanine is utilized in pre- and postoperative nutrition therapy in combination with other L-amino acids. Moreover, it serves as a sweetener in the food industry due to its pleasant taste. Over the years, the expanding industrial and scientific landscape, coupled with decreasing costs, has led to a surge in the utilization of L-Alanine. Particularly noteworthy is its role as a key raw material for the production of methylglycinediacetic acid (MGDA). MGDA, a novel synthetic green chelating agent with exceptional performance, finds applications in various cleaning products. The enhanced affordability of L-Alanine has significantly accelerated the development and adoption of MGDA. Furthermore, L-Alanine holds promise in the creation of biodegradable and biocompatible polymers like polyesteramide, as well as in the development of engineered thermoplastics such as polyamides and poly(amide-ester-imide)s. These advancements underscore the significance of L-Alanine as a versatile compound with multifaceted applications across diverse industries. ¹

Production by Engineered Microorganisms

The production of L-alanine through metabolic engineering of microorganisms has been a significant area of research since the 1990s. Various strategies have been employed to enhance the yield and efficiency of L-alanine production using engineered strains. One key approach involves the manipulation of the alanine racemase enzyme (L-ALD) in host microorganisms. Many native strains lack this enzyme, necessitating its introduction for efficient L-alanine production. By overexpressing genes such as B. sphaericus L-ALD alaD in strains like Zymomonas mobilis CP4thi or E. coli AL1 and C. glutamicum AL107, researchers have successfully enhanced L-alanine production levels. This genetic modification has led to significant improvements in L-alanine yields, with strains like C. glutamicum AL107 achieving impressive titer levels and high optical purities. Another critical aspect of optimizing L-alanine production is the minimization of coproduct formation through the deletion of genes associated with competitive metabolic pathways. By eliminating genes like ldhA and aceF in strains like Lactococcus lactis or E. coli AL887, researchers have been able to enhance L-alanine yields and purity. Additionally, further genetic engineering efforts in E. coli have shown promising results, with strains producing high levels of L-alanine approaching theoretical maximum yields. To address the challenge of L-alanine inhibition on cell growth, novel genetic switches have been designed. For instance, a thermoregulated genetic switch was implemented in an engineered E. coli strain to dynamically regulate the expression of the Geobacillus stearothermophilus alaD gene. This innovative approach has allowed for precise control over L-alanine production, resulting in significantly increased titers and yields of L-alanine, even under oxygen-deprived conditions. Overall, the continuous advancement in metabolic engineering techniques, gene manipulation, and strain optimization has significantly enhanced the production of L-alanine by engineered microorganisms. These innovative strategies not only improve yields and purity but also pave the way for more efficient and sustainable production processes in the biotechnology industry. ²

Reference

1. Bonillo Martínez AD, Galán ICR, Bellver MVM. Application of a Biodegradable Polyesteramide Derived from L-Alanine as Novel Excipient for Controlled Release Matrix Tablets. AAPS PharmSciTech. 2017; 18(8): 3286-3295.

2. Liu P, Xu H, Zhang X. Metabolic engineering of microorganisms for L-alanine production. J Ind Microbiol Biotechnol. 2022; 49(2): kuab057.