Harnessing biocompatible chemistry for developing improved and novel microbial cell factories

Abstract

White biotechnology relies on the sophisticated chemical machinery inside living cells for producing a broad range of useful compounds in a sustainable and environmentally friendly way. However, despite the impressive repertoire of compounds that can be generated using white biotechnology, this approach cannot currently fully replace traditional chemical production, often relying on petroleum as a raw material. One challenge is the limited number of chemical transformations taking place in living organisms. Biocompatible chemistry, that is non-enzymatic chemical reactions taking place under mild conditions compatible with living organisms, could provide a solution. Biocompatible chemistry is not a novel invention, and has since long been used by living organisms. Examples include Fenton chemistry, used by microorganisms for degrading plant materials, and manganese or ketoacids dependent chemistry used for detoxifying reactive oxygen species. However, harnessing biocompatible chemistry for expanding the chemical repertoire of living cells is a relatively novel approach within white biotechnology, and it could potentially be used for producing valuable compounds which living organisms otherwise are not able to generate. In this mini review, we discuss such applications of biocompatible chemistry, and clarify the potential that lies in using biocompatible chemistry in conjunction with metabolically engineered cell factories for cheap substrate utilization, improved cell physiology, efficient pathway construction and novel chemicals production.

Figure 1

Harnessing biocompatible chemistry to promote growth. 2,5 dimethoxy‐1,4‐benzoquinone (2,5‐DMBQ) and involution are secreted by certain fungi, and stimulate Fenton chemistry which results in formation of highly reactive hydroxyl radicals (^·OH) that can be used to degrade lignin.

Figure 2

The integration of an electrochemical reaction with cellular metabolism for CO₂ fixation. A. Formate is generated electrochemically from CO₂ and H₂O in the presence of Li catalyst. Formate is subsequently converted to CO₂ and NADH by formate dehydrogenase (FDH) using an engineered Ralstonia eutropha strain. NADH is used for generating ATP via oxidative phosphorylation and NADPH through transhydrogenase, two compounds that are used to drive CO₂ fixation to produce longer chain alcohols. B. The reductive glycine pathway. CBB, calvin‐benson‐bassham; FDH, formate dehydrogenase; PHB, polyhydroxybutyrate; THF, tetrahydrofolate; Tr, transhydrogenase.

Figure 3

Proposed mechanism for Mn²⁺‐dependent and ketoacid‐dependent detoxification of ROS and. A. A Mn (II) complex, such as Mn²⁺‐P (phosphate or polyphosphate), can be oxidized by ${\text{O}}_2^{-}$ to a Mn (III) complex (Mn³⁺‐P) with the generation of H₂O₂, whereafter ${\text{O}}_2^{-}$ reduces Mn (III) to Mn (II) resulting in formation of O₂. The superoxide dismutase (SOD) enzymatically can perform a similar detoxification of ROS. B. The ketoacids, such as glyoxylate, pyruvate and α‐ketoglutarate can non‐enzymatically be converted into corresponding organic acids formate, acetate and succinate in the presence of H₂O₂.

Figure 4

Producing unique chemicals by harnessing biocompatible chemistry. The biocompatible reactions involved are highlighted by green background. A. The non‐enzymatic oxidative decarboxylation from α‐acetolactate to diacetyl (Module II) links the two metabolic pathways (Module I: from glucose to α‐acetolactate by glycolysis and α‐acetolactate synthase; Module III: from diacetyl to S‐acetoin and further to (S,S)‐2,3‐butanediol by butanediol dehydrogenase). The synthetic pathway for (S,S)‐2,3‐butanediol is redox neutral. B. The Pictet–Spengler chemical reaction is involved in the condensation of dopamine and 3,4‐DHPAA (3,4‐dihydroxyphenylacetaldehyde) to form (S)‐norlaudanosoline, which is an essential precursor for the synthesis of thebaine. C. The non‐enzymatic chemical reactions are responsible for the final steps in the synthesis of artemisinin from dihydroartemisinic acid (DHAA). D. Non‐enzymatic oxygenated metabolites in the presence of reactive oxygen species, 5‐F_3t‐isoprostanes (IsoP) from EPA and F_4t‐neuroprostanes (NeuroP) from DHA. E. The production of norisoprenoid by combining enzymatic and non‐enzymatic reactions in Bacillus cells. Menaquinone‐7, which is synthesized by combination of heptaprenyl diphosphate and 1,4‐dihydroxy‐2‐naphtoate (DHNA) through enzymatic reactions, can react with superoxide non‐enzymatically to form farnesylfarnesylacetone, one non‐C5‐units terpenoid. Another two norisoprenoids farnesylacetone and phytone can be formed similarly from menaquinone‐4 and phylloquinone, respectively. F. In the presence of phthalocyanine catalyst, phenyl cyclopropane can be achieved from styrene, which is biosynthesized in metabolically engineered E. coli. The vitamin E‐derived micelles provide compartmentalized space to accommodate this chemical reaction to alleviate the toxicity of styrene and further increase the production of phenyl cyclopropane with high titre and productivity. G. Non‐enzymatic glycosylation between an aldehyde group in glucose and an amino group in para‐aminobenzoate (PABA). It was found as a byproduct for producing PABA in engineered C. glutamicum.