Gas fermentation with recombinant Cupriavidus necator for lactic acid production and PLA production

Introduction:

Polylactide (PLA) is recognized for its high mechanical strength, rigidity, effective barrier properties, and moderate thermostability, rendering it a well-suited biopolymer for applications demanding structural robustness and stability. A critical precursor for PLA synthesis is lactic acid (L- or D-isomer) and a high optical purity (≥ 99%) is essential to achieving the desired polymer quality¹. The use of chemolithoautotrophic bacterium Cupriavidus necator presents a sustainable approach for lactic acid production through gas fermentation, using CO₂ as the sole carbon source.

Goals:

This study aims to produce optically pure D- and L-lactate in C. necator, and to eventually produce a biopolymer co-polymerized from L-lactate with 3-hydroxybutyrate(3HB). Throughout the study, we will assess the potential of gas fermentation as a strategy for biopolymer synthesis, emphasizing its potential to reduce reliance on conventional feedstocks while supporting a circular bioeconomy.

Materials & Methods:

To enable reduction of pyruvate to D- or L-lactate, a D-lactate dehydrogenase (D-LDH) from Leuconostoc mesenteroides and three L-LDHs from Bos taurus, Lactobacillus helveticus, and Lactiplantibacillus plantarum were selected to be produced in C. necator H16 and PHB-synthesis deletion strains. The genes responsible for converting D-lactate or L-lactate into pyruvate were targeted for knockout using a pLO3 plasmid-based double-crossover homologous recombination to prevent reverse conversion and enhance lactate production.

Results:

To date, D-LDH from L. mesenteroides was heterologously produced in C. necator H16 wildtype and PHB-synthesis deletion mutant showing by immunoblot.

Summary:

With the successful heterologously production of D-LDH from L. mesenteroides, this study presents the potential of C. necator for sustainable lactic acid production via gas fermentation. This lays the groundwork for developing C. necator as an autotrophic biotechnological platform for PLA and its copolymer synthesis. Further strain engineering is required to maximize the yield while minimizing metabolic burden on the organism.

Reference:

1 Huang S, Xue Y, Yu B, Wang L, Zhou C, Ma Y. Molecules. 2021;26(21):6446.