Background The synthesis of 2-hydroxyisobutyric acid (2-HIB), a promising building block for, e.g., Plexiglas® production, is described as an example for a clean and sustainable bioproduction. Methods A derivative strain of Cupriavidus necator H16, impaired in the poly-ß-hydroxybutyrate synthesis pathway and equipped with xenogenic 2-hydroxyisobutyryl-coenzyme A mutase from Aquincola tertiaricarbonis L108, was applied. Batch cultivation was performed in the presence of vitamin B12 by supplying a gas mixture comprising hydrogen, oxygen, and carbon dioxide. Results Exploiting the chemo-litho-autotrophic potential of this so-called knallgas bacterium, 2-HIB was synthesized and excreted into the cultivation broth under aerobic conditions when inorganic nitrogen-limited conditions allowed an overflow metabolism of carbon metabolites. 2-HIB synthesis proceeded at a rate of 8.58 mg/[(g bacterial dry mass)·h]. Approximately 400 mg/L in total was obtained. The results were subsequently compared to calculated model data to evaluate the efficiency of the conversion of the substrates into the product. To achieve overall yield data regarding the substrate conversion, the model describes an integral process which includes both 2-HIB synthesis and biomass formation. Conclusions This study has confirmed the feasibility of the microbial synthesis of the bulk chemical 2-HIB from hydrogen and carbon dioxide by exploiting the chemo-litho-autotrophic metabolism of C. necator H16 PHB−4, additionally expressing the foreign 2-HIB-coenzyme A mutase. The product synthesis was satisfying as a proof of principle but does not yet approach the maximum value as derived from the model data. Furthermore, the biosynthesis potential of an optimized process is discussed in view of its technical application.

The maximum growth rate of methylotrophic yeasts during growth on methanol is about 0.2 h−1. Since they are able to grow faster on substrates such as glucose we tried to identify the putative limiting step in methanol metabolism within the assimilatory pathway, leading to the formation of a major precursor for biosyntheses, or within the linear dissimilatory sequence. Growth experiments with mixed substrates and determination of some kinetic parameters allowed us to restrict the number of possible pacemaker enzymes. The dissimilatory sequence does not seem to be growth‐rate limiting. This also applies to transketolase, transaldolase and fructose‐1,6‐bisphosphatase. Surprisingly, methanol oxidase appears to be the prime candidate.