Frances H. Arnold, California Institute of Technology, USA

Chemistry encoded in DNA and optimized by evolution promises efficient, clean, sustainable routes to fuels, chemicals, materials, foods, medicines, and more. Evolution not only optimizes and tunes features such as activity or stereoselectivity—it also innovates. We are using evolution to create entirely new biocatalysts that catalyze reactions unknown in biology and sometimes unprecedented in human-invented chemistry. New-to-nature ‘carbene transferase’ and ‘nitrene transferase’ enzymes increase the scope of molecules and materials that can be made using biology’s remarkable chemical machinery. Such enzymes unlock chemicial transformations that were inaccessible to small-molecule catalysts. And, with modern machine learning and AI tools to aid discovery and optimization, we are closer than ever to encoding a vast array of chemical transformations in DNA.

Marc Fontecave, Collège de France, FR

The energy transition requires new sources of carbon, as alternatives to fossil sources, for the production of organic compounds for the chemical industry (fuels, polymers, etc.). Carbon dioxide transrformation to C1 or C2 compounds can be achieved by electroreduction or photoreduction using low-carbon electricity or sunlight directly, respectively. The reactions at work imply multiple electrons and protons and thus require catalysts for minimizing energy barriers and controlling selectivity. Whereas synthetic coordination complexes and solid metal materials are extensively studied, it is interesting to note that living organisms have evolved complex metalloenzymes for the reduction of CO₂ into formic acid and for the interconversion between CO₂ and CO, providing a source of inspiration for the design of biomimetic homogeneous and heterogeneous catalysts. Another class of bioinspired catalysts, namely artificial enzymes, deserves more attention. Such a strategy has been very little explored so far for CO₂ electro- or photo-reduction. This the theme of this presentation which will focus on our recent studies of artificial CO₂ reductases.

Anthony Green, University of Manchester, UK

Protein cavities can offer highly versatile and engineerable environments for hosting new catalytic sites. However, only a narrow range of functional elements are available to enzyme designers when building new active sites, meaning that many important modes of reactivity are not accessible. Here I will discuss our efforts to overcome these limitations, by encoding new catalytic elements into proteins as non-canonical amino acid side chains. This approach has allowed us to build enzymes with new functions and reactivity modes that were previously inaccessible with protein catalysts. Significantly as our catalyts are genetically encoded, their activities and selectivities can be optimized using directed evoution workflows adpated to an expanded amino acid alphabet. We are optimistic that this integration of enzyme design, genetic code expansion and laboratory evolution can provide a versatile strategy for creating enzymes with catalytic functions not accessible to nature.

John F. Hartwig, University of California, Berkeley, USA

The introduction of functional groups at the positions of typically unreactive C-H bonds site-selectively and the stereo- and regio-selective functionalization of unconjugated C=C bonds have been longstanding challenges in catalysis. To this end, our group has been motivated by the limits of small-molecule catalysts for such reactions to begin to investigate artificial metalloenzymes. These artificial metalloenzymes contain synthetic cofactors possessing abiotic metal centers that catalyze unnatural reactions with control over selectivity resulting from the protein environment. In the best-case scenario such reactions could occur within the cells of E. coli or other microorganisms and in the long-term to occur as part of an unnatural biosynthetic pathway to produce unnatural products by fermentation.

This talk will include results on new transformations, new mechanisms, new reactive intermediates, and new methods for in vivo assembly of artificial metalloenzymes. This combination of results has enabled us to combine an unnatural carbene-transfer reaction catalyzed by natural and artificial metalloenzymes with the biosynthesis of diazo compounds and natural reactions of a heterologous biosynthetic pathway to create engineered microorganisms that produce unnatural products by artificial biosynthesis encompassing organometallic chemistry.

Xiongyi Huang, Johns Hopkins University, USA

Repurposing natural enzymes to catalyze synthetic transformations absent in nature has emerged as a significant research field bridging chemistry and biology. A key challenge in this pursuit is the introduction of synthetic reaction mechanisms into natural protein scaffolds. Over the past decades, substantial breakthroughs have been achieved in this field, with many enzymatic systems developed to catalyze critical chemical transformations not previously observed in biology. However, much of this progress has focused on proteins or enzymes containing heme or organic cofactors. In this context, our group has drawn inspiration from mechanistic connections between synthetic and biocatalytic systems to explore the vast, untapped potential of nonheme enzymes for new-to-nature biocatalysis. This talk will highlight several enzymatic systems developed by our group over the past five years, which utilize diverse reaction mechanisms in transition metal catalysis for the formation of C‒N, C‒S, C‒C, C‒O, and C‒halogen bonds. We hope these systems will further advance the integration of synthetic chemistry and biology to innovate chemical synthesis, as well as deepen our understanding of both biochemical and synthetic reaction mechanisms.

Todd Hyster, Princeton University, USA

Enzymes are exquisite catalysts for chemical synthesis, capable of providing unparalleled levels of chemo-, regio-, diastereo- and enantioselectivity. Unfortunately, biocatalysts are often limited to the reactivity patterns found in nature. In this talk, I will share my groups efforts to use light to expand the reactivity profile of enzymes. In our studies, we have developed novel photoexcitation mechanisms involving common biological cofactors, such as nicotinamide, flavin, and pyridoxal, to facilitate electron transfer to substrates bound within enzyme active sites. Alternatively, proteins can be used to electronically activate substrates for reduction by exogenous photoredox catalysts enabling radical formation to be localized to the protein active site. The resulting radicals can engage in a variety of inter- and intramolecular reactions with high levels of enantioselectivity. These approaches enable biocatalysts to solve long-standing selectivity challenges in chemical synthesis.

Yi Lu, University of Texas, USA

Metalloenzymes play important roles in many biological processes, yet the structural features underlying their remarkable reactivity and selectivity remain incompletely understood. To address this issue, we have designed artificial metalloenzymes (ArMs) using small, stable proteins as scaffolds. These scaffolds are designed to incorporate key residues essential for functions in native enzymes, including O₂ and N₂ reduction as well as nitrosylation and polysaccharide oxidative cleavage reactions.

Our findings reveal that replicating the primary coordination sphere may suffice for creating structural models of metalloenzymes. However, achieving functional ArMs with high activity and turnover rates—on par with native enzymes—requires precise engineering of non-covalent secondary coordination sphere interactions, such as hydrophobic effects and hydrogen bonding, including those mediated by water molecules.

This presentation will highlight recent advances in ArM design, the insights gained from these studies, and the expansion of ArM activities beyond the capabilities of natural enzymes.

Paolo Melchiorre, University of Bologna, IT

The combination of photocatalysis, biocatalysis, and organocatalysis provides a powerful yet underexplored strategy for addressing major challenges in asymmetric synthesis. By combining these distinct catalytic disciplines, we aim to develop novel enantioselective radical processes that are otherwise difficult to achieve using conventional approaches. Central to this concept is the ability to harness enzyme-bound organocatalytic intermediates as photoactive intermediates, unlocking new mechanistic pathways for radical generation and control.

Recently, we developed a new approach to light-driven biocatalysis, where engineered enzymes utilize iminium ion intermediates—formed transiently within their active sites—as single-electron oxidants upon visible-light excitation. This strategy enables the activation of chiral carboxylic acids, triggering radical decarboxylation and subsequent stereospecific cross-coupling to construct complex chiral architectures with multiple stereocenters and complete enantiocontrol. Notably, the enzyme’s active site prevents racemization of chiral radicals via a rare "memory of chirality" mechanism, ensuring high stereochemical fidelity. By leveraging the unique synergy between light, biocatalysts, and organocatalytic intermediates, this work expands the scope of radical chemistry and sets the stage for new, sustainable methods in asymmetric synthesis.

Hannah S. Shafaat, University of California, Los Angeles, USA

Metalloenzymes catalyze the challenging chemical reactions that lie at the core of vital life processes, from carbon and nitrogen fixation to photosynthesis and respiration. Nickel-containing enzymes, specifically, are essential for global hydrogen and carbon cycling and the metabolisms of diverse microbes, with implications in human health, clean energy conversion, and sustainable fuel generation. In this presentation, I will discuss our recent efforts to recapitulate key structural and functional elements of microbial nickel enzymes such as hydrogenase, carbon monoxide dehydrogenase (CODH), and acetyl coenzyme A synthase using protein-based scaffolds. By designing model metalloenzymes from the “inside out”, each contribution can be clearly delineated. Functional studies of our model proteins are combined with diverse spectroscopic techniques and computational investigations, allowing us to obtain a comprehensive understanding of how the entire protein matrix contributes to reactivity. These fundamental structure-function-dynamics relationships will be discussed in the context of understanding native metalloenzymes and providing design guidelines for new biological and anthropogenic catalyst development.

Woon Ju Song, Seoul National University, KR

Protein scaffolds offer an expansive platform to construct novel structures and functions of metallocofactors. While numerous methods have been developed to optimize protein environments near or distant from active sites, the design of the first coordination sphere still relies largely on structure-based approaches derived from coordination chemistry. Here, we present two strategies to build first coordination spheres for new functions: (i) pinpointing specific positions for mutations to create divalent transition metal-binding sites by developing an in silico program, Metal-Installer, and (ii) genetically incorporating noncanonical amino acids followed by symmetry-based ligand mulitplication. By integrating geometric parameters derived from both natural metalloproteins and synthetic inorganic complexes, we created tailor-made metal-binding sites that mimic the structure and chemical properties of mono- and dinuclear metalloproteins. These designs closely matched our structural predictions and fulfilled the minimal requirements for metal-dependent catalysis or photochemical properties. This work significantly broadens the accessible chemical space of metalloproteins, enabling the repurposing of natural protein/enzyme scaffolds for various applications such as metalloenzyme mimics, biocatalysts, and protein-based photochemical materials.

Yuzhou Wu, Huazhong University of Science and Technology, CN

Biomanufacturing is a transformative technology with the potential to revolutionize diverse industrial sectors. An underlying limitation in this field is the lack of enzymes capable of driving unnatural transformations necessary for the synthesis of a broad range of chemical products. However, enzyme design is inherently restricted by the availability of structural and functional units provided by nature. This presentation will detail our ongoing efforts to develop new enzymes that incorporate unnatural entities into their scaffolds, with a particular emphasis on photoenzymes that utilize synthetic photosensitizers. I will discuss strategies for incorporating these entities, including genetic encoding approaches and chemical modification techniques, and studying their compatibility with high-throughput directed evolution in living cells. Additionally, I will highlight the unique catalytic properties observed in these artificial enzymes and their potential to be integrated with natural cellular metabolic pathways. We envision that artificial enzymes, enriched with unnatural entities, will serve as versatile tools for constructing "cell factories" adaptable to a variety of synthetic applications.

Nico Bruns, Technical University of Darmstadt, DE

Atom transfer radical reactions are chemical transformations that do not occur naturally in biological systems. Yet, metalloenzymes can catalyze such new-to-nature reactions, thereby paving the way to biocatalytic routes to atom transfer radical polymerizations (bioATRP) and atom transfer radical cyclizations (bioATRC). However, native heme proteins such as myoglobin and horseradish peroxidase display a limited degree of control or activity in these reactions. By rational design, myoglobin mutants that show enhanced performance over their wild-type counterparts in radical polymerizations and greatly enhanced catalytic turnover in radical cyclization reactions were created. Moreover, bioATRP is not only an enzymatic route to polymers but allows the synthesis of polymers in situ in biological systems. This opens up the possibility to engineer living cells on their surface and within their cytosol by biocatalytic radical polymerizations and to create artificial cells that can express their own proteins. Thus, repurposed metalloenzymes play a crucial role in developing novel communicating life-like biomaterials, semi-synthetic engineered living materials, and synthetic biology systems.

Jared C. Lewis, Indiana University, USA

Metalloenzymes perform some of the most remarkable transformations in nature under ambient conditions in complex cellular milieu. The possibility of leveraging molecular recognition and evolution for non-biological metal catalysts has driven efforts to engineer artificial metalloenzymes (ArMs), hybrid catalysts comprised of synthetic metal cofactors linked to protein scaffolds. In this talk, I will discuss recent efforts from my group to design and evolve ArMs containing dirhodium and metal polypyridine cofactors for selective catalysis.

Angela Lombardi, University of Naples Federico II, IT

Metalloenzymes are capable of catalyzing a variety of reactions, and a given metal ion can be used in a number of oxidative, reductive, and hydrolytic transformations in different enzymes. This functional diversity arises from a strong partnership between the metal cofactor and protein matrix: the metal ion provides the protein with an array of chemical properties, while the protein stabilizes it in solution and directs its reactivity toward a unique and distinct path.

Bioinorganic chemists tackled the challenge to unravel the mechanisms that allow the protein matrix to modulate the catalytic activity of metal-containing cofactors, through the development of artificial systems. In this respect, de novo protein design, involving the construction of proteins “from scratch”, has contributed to tremendous advances in manufacturing metalloenzymes with unique structures and functionalities.

This lecture will give an overview of our results on the design of artificial helical bundles, housing different metal cofactors, which catalyze a variety of oxidative reactions. Starting from the Due Ferri (DF) family of artificial diiron-oxo-proteins, our design has integrated rational and computational strategies to engineer mononuclear and dinuclear copper sites, mimicking natural lytic polysaccharide monooxygenases (LPMOs) and polyphenol oxidases.

Osami Shoji, Nagoya University, JP

Cytochrome P450BM3 is a highly efficient heme enzyme, but its native activity is largely restricted to long-chain fatty acids. To overcome this limitation without relying on mutagenesis, we developed a decoy molecule strategy that employs inert compounds mimicking native substrates to trigger oxygen activation. This approach enabled wild-type P450BM3 to hydroxylate abiological substrates such as benzene, propane, and methane. Optimized decoy molecules, particularly those derived from N-acyl amino acids and amino acid dimers, enhanced substrate turnover and, in some cases, promoted crystallization of the enzyme, which facilitated structural analysis. This strategy has also been successfully applied to other P450 enzymes, demonstrating its broader applicability. In addition, by combining the decoy approach with directed evolution, we developed variants capable of utilizing microbial signaling molecules such as N-acyl homoserine lactones as functional decoys. Our decoy molecule strategy offers new opportunities for sustainable and programmable oxidation chemistry.

Cathleen Zeymer, Technical University of Munich, DE

Cerium photoredox catalysis is a powerful method to activate organic molecules under mild conditions. However, it remains a major challenge to achieve stereocontrol in these light-driven radical reactions. We thus developed a cerium-dependent photoenzyme enabling this chemistry in the chiral environment of a de novo protein. Our work is based on a de novo TIM barrel scaffold designed previously in a physics-based approach. We equipped the protein with a high-affinity metal binding site for lanthanide ions and demonstrated its photocatalytic potential. Upon visible-light irradiation, the cerium-bound enzyme enables the radical C–C bond cleavage of 1,2-diols in aqueous solution. To optimize the initially low activity and enantioselectivity, we redesigned the scaffold computationally. We decreased the cavity size between the two independently folded domains and used ProteinMPNN to redesign the sequences. Selected redesigns were characterized experimentally and showed significantly improved k_cat/K_M and enantiomeric excess in kinetic resolutions of diols.

Huimin Zhao, University of Illinois Urbana-Champaign, USA

Enzymes have been increasingly used for practical synthesis of chemicals, fuels, and materials thanks to recent advances in enzyme engineering, synthetic biology, artificial intelligence (AI)/machine learning (ML), and laboratory automation. In this talk, I will discuss our recent effort in designing repurposed enzymes with new-to-nature reactivity for asymmetric synthesis by exploring the synergy between enzymatic catalysis and photocatalysis. The representative new-to-nature photoenzymatic reactions that we have demonstrated so far include but are not limited to intermolecular radical hydroalkylation, intermolecular radical conjugate addition, and intermolecular radical hydroamination. In addition, I will introduce a new strategy to address the scalability issue of these new-to-nature photoenzymatic reactions by directly integrating them into microbial metabolism. Finally, I will highlight the development of machine learning and laboratory automation tools for enzyme discovery and engineering. Taken together, these strategies and tools should greatly accelerate the development of biocatalysts for applications related to human health, energy, and sustainability.