title: Exploiting mixtures of H2, CO2, and O2 for improved production of methacrylate precursor 2-hydroxyisobutyric acid by engineered Cupriavidus necator strains.
abstract: Current manufacturing of most bulk chemicals through petrochemical routes considerably contributes to common concerns over the depletion of fossil carbon sources and greenhouse gas emissions. Sustainable future production of commodities thus requires the shift to renewable feedstocks in combination with established or newly developed synthesis routes. In this study, the potential of Cupriavidus necator H16 for autotrophic synthesis of the building block chemical 2-hydroxyisobutyric acid (2-HIBA) is evaluated. A novel biosynthetic pathway was implemented by heterologous expression of the 2-hydroxyisobutyryl-coenzyme A (2-HIB-CoA) mutase from Aquincola tertiaricarbonis L108, relying on a main intermediate of strain H16's C4 overflow metabolism, 3-hydroxybutyryl-CoA. The intention was to direct the latter to 2-HIBA instead or in addition to poly-3-hydroxybutyrate (PHB). Autotrophic growth and 2-HIBA (respectively, PHB) synthesis of wild-type and PHB-negative mutant strains were investigated producing maximum 2-HIBA titers of 3.2 g L(-1) and maximum specific 2-HIBA synthesis rates (q 2-HIBA) of about 16 and 175 μmol g(-1) h(-1), respectively. The obtained specific productivity was the highest reported to date for mutase-dependent 2-HIBA synthesis from heterotrophic and autotrophic substrates. Furthermore, expression of a G protein chaperone (MeaH) in addition to the 2-HIB-CoA mutase subunits yielded improved productivity. Analyzing the inhibition of growth and product synthesis due to substrate availability and product accumulation revealed a strong influence of 2-HIBA, when cells were cultivated at high titers. Nevertheless, the presented results imply that at the time the autotrophic synthesis route is superior to thus far established heterotrophic routes for production of 2-HIBA with C. necator.
authors.affiliation: Department of Environmental Microbiology, Helmholtz Centre for Environmental Research UFZ, Permoserstrasse 15, 04318, Leipzig, Germany, deniseprz46@gmail.com.
authors.forename: Denise
authors.initials: D
authors.lastname: Przybylski
authors.forename: Thore
authors.initials: T
authors.lastname: Rohwerder
authors.forename: Cornelia
authors.initials: C
authors.lastname: Dilßner
authors.forename: Thomas
authors.initials: T
authors.lastname: Maskow
authors.forename: Hauke
authors.initials: H
authors.lastname: Harms
authors.forename: Roland H
authors.initials: RH
authors.lastname: Müller
mesh_terms: D053898:Biosynthetic Pathways / Q000235:genetics; D002245:Carbon Dioxide / Q000378:metabolism*; D020118:Cupriavidus necator / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D006859:Hydrogen / Q000378:metabolism*; D006885:Hydroxybutyrates / Q000378:metabolism*; D019751:Intramolecular Transferases / Q000235:genetics / Q000378:metabolism*; D060847:Metabolic Engineering; D010100:Oxygen / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D006885:Hydroxybutyrates; D011994:Recombinant Proteins; D002245:Carbon Dioxide; D006859:Hydrogen; C008039:2-hydroxyisobutyric acid; D019751:Intramolecular Transferases; D010100:Oxygen