Carboxylases in natural and synthetic microbial pathways

Abstract

Carboxylases are among the most important enzymes in the biosphere, because they catalyze a key reaction in the global carbon cycle: the fixation of inorganic carbon (CO₂). This minireview discusses the physiological roles of carboxylases in different microbial pathways that range from autotrophy, carbon assimilation, and anaplerosis to biosynthetic and redox-balancing functions. In addition, the current and possible future uses of carboxylation reactions in synthetic biology are discussed. Such uses include the possible transformation of the greenhouse gas carbon dioxide into value-added compounds and the production of novel antibiotics.

Fig. 1.

Physiological classification of carboxylases. The scheme illustrates the five different physiological functions of carboxylases defined in this article. Autotrophic carboxylases serve in autotrophic pathways that allow the formation of central precursor molecules (PEP, pyruvate, acetyl-CoA, and TCA cycle intermediates) solely from inorganic carbon (CO₂). Assimilatory carboxylases function in heterotrophic pathways that convert an organic growth substrate into central precursor molecules. Anaplerotic carboxylases are involved in refilling TCA cycle intermediates when those are drained for biosynthesis. Biosynthetic carboxylases operate in the synthesis of cellular building blocks from central precursor molecules (e.g., α-carboxylic thioesters for fatty acid and polyketide biosynthesis). Redox-balancing carboxylases operate in the transfer of reducing equivalents onto carbon dioxide as terminal electron acceptors and are important for redox homeostasis.

Fig. 2.

Autotrophic carboxylation reactions. All carboxylases used in autotrophic CO₂ fixation pathways are listed and sorted according to “efficiency.” Carboxylases that do not require ATP hydrolysis for their carboxylation reaction are considered to be more efficient (“energetic efficiency”). Similarly, carboxylases that combine carboxylation with reduction, or allow a subsequent reduction step without extra ATP hydrolysis, are also considered to be more efficient (“catalytic power”). Note that carboxylase efficiency is directly linked to the total energetic costs of a given pathway (see the text for a detailed discussion).

Fig. 3.

Assimilatory carboxylation reactions I: substrate functionalization of inert substrates by carboxylation. Many compounds that bear no functional terminal group are assimilated through an initial carboxylation step. The carboxylic acid formed is then transformed into a CoA ester and further channeled into central metabolites (e.g., by classical β-oxidation). Functionalization of substrates by carboxylation is a common strategy under anaerobic conditions, when no molecular oxygen is present to activate the substrate.

Fig. 4.

Assimilatory carboxylation reactions II: propionate, leucine and acetate assimilation. Many compounds are initially channeled into few CoA esters (e.g., propionyl-CoA, acetyl-CoA) during assimilation. The further conversion of these CoA esters into central precursor molecules requires carboxylation. Acetate is transformed initially into acetyl-CoA and can be assimilated via three different carboxylative routes (pyruvate:ferredoxin oxidoreductase [circled “1”]; ethylmalonyl-CoA pathway [circled “2”]; the methylaspartate cycle [circled “3”]. Note the importance of propionyl-CoA carboxylation in many of the assimilation pathways (highlighted in yellow).