Organizing Multi-Enzyme Systems into Programmable Materials for Biocatalysis

Abstract

Significant advances in enzyme discovery, protein and reaction engineering have transformed biocatalysis into a viable technology for the industrial scale manufacturing of chemicals. Multi-enzyme catalysis has emerged as a new frontier for the synthesis of complex chemicals. However, the in vitro operation of multiple enzymes simultaneously in one vessel poses challenges that require new strategies for increasing the operational performance of enzymatic cascade reactions. Chief among those strategies is enzyme co-immobilization. This review will explore how advances in synthetic biology and protein engineering have led to bioinspired co-localization strategies for the scaffolding and compartmentalization of enzymes. Emphasis will be placed on genetically encoded co-localization mechanisms as platforms for future autonomously self-organizing biocatalytic systems. Such genetically programmable systems could be produced by cell factories or emerging cell-free systems. Challenges and opportunities towards self-assembling, multifunctional biocatalytic materials will be discussed.

Facile Synthesis of Sugar Nucleotides from Common Sugars by the Cascade Conversion Strategy

“…Although de novo biosynthesis undergoes complex reactions as mentioned above, the last bioconversion reactions are often oxidation, reduction, amination, and acetylation reactions, and these reactions require cofactors such as NADH/NADPH, NAD + /NADP + , pyridoxamine-5′-phosphate (PMP), or acetyl coenzyme A (AcCoA). , Multistep enzymatic reactions can be performed in one-pot due to the high stereo- and region-selectivity of enzyme-catalyzed reactions, which is also known as the OPME (one-pot multienzyme) synthetic strategy. − Thus, we reorganized complex de novo biosynthesis as a cascade reaction to prepare target sugar nucleotides, aiming to make the reactions proceed cleanly. If the reactions could proceed cleanly, complex de novo bioconversions will become practical for synthetic use.…”

Confining Enzyme Clusters in Bacteriophage P22 Enhances Cofactor Recycling and Stereoselectivity for Chiral Alcohol Synthesis

“…In contrast, living organisms maximize the efficiency of cofactor recycling based on the specific spatial organization of enzymes in confined cellular environments . Inspired by nature, many artificial nanostructures have been created to increase the efficiency of enzyme-catalyzed reactions occurring in vitro. ,, Recently, virus-like particles (VLPs) have been engineered as nanoreactors to encapsulate single or multiple enzymes to perform enzymatic catalysis . Hydrogen production has been achieved in P22 nanoreactors using complex [NiFe]- or [FeFe]-hydrogenases. , In addition, enzymes have also been confined in nanoscale microbial compartments including carboxysomes and encapsulins. , However, to date, few reports have examined the feasibility of cofactor recycling using VLP-encapsulated enzymes for the synthesis of chiral chemicals.…”

Extracellular Electron Transfer Enables Cellular Control of Cu(I)-Catalyzed Alkyne–Azide Cycloaddition

“…Biological catalysis provides several advantages over traditional chemical catalysis including milder operating conditions, self-regeneration, and the ability to optimize activity via genetic manipulation. − Whole-cell biocatalysts can leverage additional dynamic control over such reactions by coupling activity to cellular growth and metabolism . However, reactions catalyzed by whole cells are typically limited to known metabolic transformations. , While efforts to augment the substrate scope of enzymatic reactions via directed evolution have been highly successful, there is still an ongoing need to expand the synthetic capabilities of live cells. ,, …”