Streamlining Design, Engineering, and Applications of Enzymes for Sustainable Biocatalysis

Abstract

In this Perspective we show how an expansion of the scope and impact of biocatalysis in industrial organic synthesis is enabled by streamlining the underpinning biocatalyst and bioprocess engineering. We begin by discussing how the underlying need for waste reduction and high (enantio)­selectivities fostered the introduction of biocatalysis as a sustainable technology for the industrial synthesis of active pharmaceutical ingredients (APIs). We continue by showing how advances in molecular biology, in particular gene sequencing and protein engineering, enabled the development of more and better enzymes, thereby broadening the industrial scope of biocatalysis. Further process improvements are provided through protein engineering for enzyme immobilization and integration of enzyme production with in vivo immobilization. Finally, the use of immobilized enzymes in continuous operation (biocatalysis in flow) facilitates the sequential integration of multi-step reactions into enzymatic or chemo-enzymatic cascade processes, thus enabling the complete, cost-effective, and environmentally attractive production of APIs.

Enzyme Immobilization and Co-Immobilization: Main Framework, Advances and Some Applications

“…Several thorough and comprehensive reviews focused on enzyme immobilization have been recently published, where updated information on the methodologies for enzyme immobilization and related issues, e.g., materials used for immobilization, physical and (bio)chemical characterization methods, impact on catalytic features of the enzyme and interface with reactor operation, can be found [30,36,57,67,70,[144][145][146][147][148][149][150][151][152][153][154][155].…”

“…Co-immobilization of enzymes, which act either sequentially or in a coordinated manner, involves the containment of multiple enzymes in the same space (either carrier or as CLEAs, hence denominated combi-CLEAs). This approach mimics Nature, where countless chemical reactions are catalyzed in tuned pathways within living cells [25,30,157,164,[171][172][173][174][175][176]. Co-immobilization can lead to enhanced reaction yields as it allows the optimization of several parameters for each individual nanobiocatalyst, such as size, solubility, activity, or selectivity [164,[174][175][176][177].…”

“…This latter and most relevant drawback can be overcome with enzyme Processes 2022, 10, 494 2 of 31 immobilization, an approach that decreases the overall process production economic cost, by enabling either continuous or repeated batch operation and facilitating downstream processing [24,26,27]. Moreover, immobilized enzymes typically display enhanced pH and thermal stabilities, and occasionally improved activity and selectivity, as compared to their corresponding free form counterparts [23,[28][29][30][31][32]. Additionally, by enabling continuous operation, the use of immobilized enzymes is associated with high space time yields [33].…”

Recent Developments in the Synthesis of β-Diketones

“…Biocatalysts has proven to be an excellent and greener complement for classical organic synthesis, as the employ of biocatalysts (either native, chemically, or genetically modified) allows very clean and selective procedures [97][98][99][100][101][102][103][104][105][106]. Regarding the biocatalytic synthesis of 1,3-diketones, a straightforward methodology was initially reported in 2010 by Müller and coworkers [107] by using a recombinant enzyme (YpYerE, a thiamine diphosphate (ThDP)-dependent aldolase from Yersinia pseudotuberculosis O:VI).…”

Modulating Glycoside Hydrolase Activity between Hydrolysis and Transfer Reactions Using an Evolutionary Approach

“…Enzymes are accelerators of chemical reactions that occur in living cells, which also work in vitro, making their use in the laboratory, in medical applications, and in industry possible [1][2][3]. Tailoring an enzyme's ability to carry out specific reactions is one of the greatest challenges that must be met in order to move on to a more sustainable biocatalysis process [4]. In this sense, directed evolution has proven to be a valuable strategy for evolving functions, with the limitation of requiring extensive screening efforts, in order to find an improved biocatalyst [5,6].…”