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Living systems contain a vast network of metabolic reactions, providing a wealth of enzymes and cells as potential biocatalysts for chemical processes. The properties of protein and cell biocatalysts—high selectivity, the ability to control reaction sequence and operation in environmentally benign conditions—offer approaches to produce molecules at high efficiency while lowering the cost and environmental impact of industrial chemistry. Furthermore, biocatalysis offers the opportunity to generate chemical structures and functions that may be inaccessible to chemical synthesis. Here we consider developments in enzymes, biosynthetic pathways and cellular engineering that enable their use in catalysis for new chemistry and beyond. This Review considers developments in enzymes, biosynthetic pathways and cellular engineering that enable their use in catalysis for new chemistry and beyond.

Living systems provide a promising approach to chemical synthesis, having been optimized by evolution to convert renewable carbon sources, such as glucose, into an enormous range of small molecules. However, a large number of synthetic structures can still be difficult to obtain solely from cells, such as unsubstituted hydrocarbons. In this work, we demonstrate the use of a dual cellular–heterogeneous catalytic strategy to produce olefins from glucose using a selective hydrolase to generate an activated intermediate that is readily deoxygenated. Using a new family of iterative thiolase enzymes, we genetically engineered a microbial strain that produces 4.3 ± 0.4 g l−1 of fatty acid from glucose with 86% captured as 3-hydroxyoctanoic and 3-hydroxydecanoic acids. This 3-hydroxy substituent serves as a leaving group that enables heterogeneous tandem decarboxylation–dehydration routes to olefinic products on Lewis acidic catalysts without the additional redox input required for enzymatic or chemical deoxygenation of simple fatty acids.A dual cellular-then-heterogeneous catalysis strategy has been used to produce olefins from glucose. 3-Hydroxy acids are made using an engineered microbial host. A hydrolytic step then provides the driving force for fatty acid deoxygenation by simple heterogeneous Lewis acid catalysis. This decarboxylation–dehydration route to olefinic products avoids the need for an additional redox input typically required for deoxygenation of unmodified fatty acids.

Biology holds an amazing propensity for chemistry. Living systems continuously carry out a vast plethora of chemical reactions within a complex network, known as metabolism, to sustain growth and improve evolutionary fitness. Metabolic engineers seek to utilize this aptitude for chemistry by creating biological catalysts for chemical production from renewable feedstocks. Biological catalysts offer an eco-friendly, and in some cases, superior, alternative to petrochemical-based chemical production.In this work, we examine biological catalysts designed for the production of two C4 commodity chemicals, n-butanol and (R)-1,3-butanediol. These catalysts are strains of Escherichia coli containing constructed biosynthetic pathways. Leveraging an anaerobic growth selection, laboratory adaptive evolution identified several mutant strains with improved phenotypes. We set out to understand the mechanism by which these adaptive mutations confer improved production. Through detailed analysis of n-butanol fermentation, we discovered that the parent strain for our evolution was unable to support sustained anaerobic growth via n-butanol fermentation, potentially due to metabolic burden associated with overexpression of the pathway enzymes. Further experimentation suggested that the mutations arose as a strategy to relieve metabolic burden through decreased expression of our biosynthetic pathway. The results of this study highlight the importance of balanced pathway expression when designing biological catalysts.We then shifted our focus to design a microbial catalyst for production of polyhydroxyalkanoates (PHAs) containing unsaturated monomers. Sites of unsaturation provide functional handles for downstream chemical modification. We devised a metabolic strategy to convert two non-canonical amino acids with unsaturated functional groups to their respective 2-hydroxy acids and activate these acids as coenzyme A thioesters for polymerization within E. coli. We identified and tested candidate enzymes for the appropriate activities in vitro and successfully showed that our identified enzymes can form a functional biosynthetic pathway. These experiments lay the groundwork for creation of a microbial catalyst capable of generating PHAs with unsaturated functional groups using glucose as a carbon source.

Living organisms carry out a wide range of remarkable functions, including the synthesis of thousands of simple and complex chemical structures for cellular growth and maintenance. The manipulation of this reaction network has allowed for the genetic engineering of cells for targeted chemical synthesis, but it remains challenging to alter the program underlying their fundamental chemical behavior. By taking advantage of the unique ability of living systems to use evolution to find solutions to complex problems, we have achieved ~95% theoretical yield of three C4 commodity chemicals, n-butanol, 1,3-butanediol, and 4-hydroxy-2-butanone. Genomic sequencing of the evolved strains identified pcnB and rpoBC as two gene loci that are able to alter carbon flow by remodeling the transcriptional landscape of the cell, highlighting the potential of synthetic pathways as a tool to identify metabolic control points.Competing Interest StatementThe authors have declared no competing interest.