Engineering Cupriavidus necator for L-threonine production.

L-threonine plays pivotal roles in the medicine, food, and feed industries. In this work, C. necator H16 was engineered for heterotrophic and autotrophic production of L-threonine. Blocking poly-hydroxybutyrate biosynthesis redirected the carbon flux toward L-threonine, resulting in a 6.8-fold increase in L-threonine titer relative to the wild type (from 5.56 mg/L to 38.03 mg/L). Deletion of genes involved in L-threonine catabolism further minimized L-threonine loss. Moreover, strategies including sufficient precursor supply, screening of key enzymes, relief of feedback inhibition, and efficient efflux of target products were adopted for strain modification. Overexpression of key genes in the L-threonine biosynthetic pathway successfully channeled carbon flow toward L-threonine synthesis, with peak titer reaching 102.90 mg/L. Finally, in fermenter cultivation, the maximum L-threonine titer of the best L-threonine-producing strains reached 315.40 mg/L. In autotrophic fermentation the L-threonine titer was 76.26 mg/L using CO2 as carbon source. This work provides a new way for CO2 bioconversion to L-threonine.