Functional plasticity of HCO₃⁻uptake and CO₂fixation inCupriavidus necatorH16

This is the authors' abstract. We don't add key points for this paper.

Uptake and fixation of CO 2 are central to strategies for CO 2 -based biomanufacturing. Cupriavidus necator H16 has emerged as a promising industrial host for this purpose. Despite its prominence, the ability to engineer C. necator inorganic carbon uptake and fixation is underexplored. Here, we test the role of endogenous and heterologous genes on C. necator inorganic carbon metabolism. Deletion of one of the four carbonic anhydrases in C. necator , β-carbonic anhydrase can , had the most deleterious effect on C. necator autotrophic growth. Replacement of this native uptake system with several classes of dissolved inorganic carbon (DIC) transporters from Cyanobacteria and chemolithoautotrophic bacteria recovered autotrophic growth and supported higher cell densities compared to wild-type (WT) C. necator in saturating CO 2 in batch culture. Several heterologous strains with Halothiobacillus neopolitanus DAB2 (hnDAB2) expressed from the chromosome in combination with diverse rubisco homologs grew in CO 2 equally or better than the wild-type strain. Our experiments suggest that the primary role of Can carbonic anhydrase during autotrophic growth is for bicarbonate accumulation to support anaplerotic metabolism, and an array of DIC transporters can complement this function. This work demonstrates flexibility in HCO 3 - uptake and CO 2 fixation in C. necator , providing new pathways for CO 2 -based biomanufacturing.