Research Portfolio

Overview

With over 14 years of experience in the research, my expertise ranges from chemical synthesis to advanced inorganic spectroscopy. My research has focused on uncovering the fundamental mechanisms that enable enzymes with transition metal active sites to activate some of the most chemically inert bonds. In both nature and industry, transition metals provide a versatile site for catalyzing important chemical transformations. Biological catalysts provide profound insights into how these transition metals can be deployed for industrially applications, providing inspiration for the design of both homogenous and heterogenous catalysts. Below, I relate projects I have worked on during my time at the Max Planck Institute for Chemical Energy Conversion (MPI CEC) and The University of Texas at Austin (UT Austin).

The research conducted at MPI CEC was part of an international collaboration with institutions from Germany, Norway, and Italy called The CuBE Project (https://www.cube-synergy.eu) to study C–H bond activation in nature and use these insights to develop heterogenous catalysts derived from Metal-Organic-Frameworks (MOFs) and Zeolites. During this time, I heavily employed X-ray spectroscopy, along with other inorganic spectroscopy methods to investigate Cu-based enzymes. My research at UT Austin pursued a biomimetic chemical synthesis approach to study N≡N triple bond activation and strategies for ammonia synthesis from dinitrogen. For this research, I developed a novel synthetic strategy for ligand substitution in iron-carbide clusters, laying the groundwork for the future synthetic approach of the project. Through these projects, I have worked to uncover molecular principles that can guide the development of next-generation catalysts for sustainable energy and chemical applications.

Projects:

C–H bond Hydroxylation by LPMOs

Nitrogen Fixation (N2→NH3)

C–H bond Hydroxylation by LPMOs

Investigations using X-ray Spectroscopy

Lytic polysaccharide monooxygenases (LPMOs) are copper-dependent enzymes that catalyze the oxidative cleavage of recalcitrant polysaccharides, such as chitin and cellulose, via C–H bond hydroxylation. These enzymes are not only central to biomass decomposition and the global carbon cycle but also serve as promising biocatalysts for industrial applications, such as biofuel production and green chemistry. The selective activation of strong C–H bonds is a fundamental goal in energy research, as it enables the direct conversion of abundant hydrocarbons into fuels and value-added chemicals. LPMOs activate such strong C–H bonds (~95 kcal/mol) using a single copper center coordinated by a "histidine brace" motif.

My research focuses on using primarily X-ray spectroscopy to unravel the geometric and electronic transformations that occur at the copper active site during catalysis. By studying native and mutated LPMOs in various chemical and electronic states, much insight can be gained into how LPMOs control reactivity and direct oxidative power toward productive hydroxylation pathways. These insights not only enhance our understanding of enzymatic C–H activation but also inform the design of bioinspired copper catalysts for energy and synthetic applications.

This research was done as part of an international collaboration with institutions from Germany, Norway, and Italy called The CuBE Project, funded by the European Research Council. Additionally, I was awarded support from a Humboldt Research Fellowship for this research.

(a) The LPMO metalloenzymes perform C–H oxidation at the glycosidic (C1, C4) site of insoluble polysaccharides. (b) Surface view of a chitin-active LPMO enzyme, called SmAA10A, bound to its insoluble substrate, illustrating the interfacial nature of catalysis. The Cu site resides at the periphery of the protein to interface with the substrate. (c) Close-up of the the active site of SmAA10A, which features a Cu bound to the protein by His114 and the N-terminal His28, which together make up the "histidine brace". A conserved Glu (or Gln) residue and aromatic side chains contribute to catalytic control.

Substrate Binding Effects on the Cu Site

This project focused on how substrate binding alters the electronic and structural environment of the Cu site in SmAA10A, a chitin-specific LPMO. Using a combination of X-ray Absorption Spectroscopy (XAS), X-ray Emission Spectroscopy (XES), and computational methods, we examined both Cu(II) and Cu(I) forms of SmAA10A in substrate-free and chitin-bound states. Notably, this study represents the first reported XAS/XES characterization of an LPMO in the substrate-bound state. X-ray spectroscopic methods present a notable advantage over conventional inorganic spectroscopy techniques (e.g., UV-vis, MCD), which are often incompatible with heterogenous, chitinous samples. In the particular case of the closed-shell d10 Cu(I)-containing reductively primed state, X-ray spectroscopic methods are uniquely suited for characterizing the EPR- and optically-silent Cu center.

Our results revealed that chitin binding induces significant changes in the Cu(II) site corresponding with a loss of a water ligand to accommodate the binding event. In the Cu(I) case, substrate binding-induced conformational change in Cu(I) site geometry and concurrent modulations to the electronic structure, which prime the enzyme for targeted C–H activation with an H2O2 co-substrate. The copper center adopts a more reactive coordination environment in the presence of substrate, which is hypothesized to steer the reaction toward productive peroxygenase chemistry and away from deleterious off-pathway reactions. These findings deepen our understanding of the LPMO peroxygenase mechanism and underscore the importance of protein–substrate interactions in directing oxidative chemistry.

Publications:

Joseph, C., et al. Structural and Electronic Modulations of LPMO upon Chitin Binding: Insights from X-ray Spectroscopy. Under review at J. Am. Chem. Soc.
Inorg. Chem. 2024, 63, 11063–11078. DOI: 10.1021/acs.inorgchem.4c00602

Reactivity Effects from Secondary Coordination Sphere

In this project, we explored how a single conserved second-sphere residue (glutamine or glutamate in native LPMOs) modulates the reactivity of the copper center. A fungal LPMO NcAA9C was manipulated using site-directed mutagenesis. Using spectroscopic characterization (EPR, XAS) and density functional theory (DFT), we found that substitution of the conserved Gln with Glu, Asp, or Asn affects the copper site indirectly by influencing protonation states, electron transfer rates, and reactivity toward H2O2. These findings demonstrate how the secondary sphere tunes the tunes the mechanism with which the active site proceeds. By identifying how second-sphere architecture shapes copper-mediated C–H activation, this study provides design principles for engineering synthetic catalysts for productive C–H bond activation.

J. Am. Chem. Soc. 2023, 145, 18888–18903. DOI: 10.1021/jacs.3c05342

Nitrogen Fixation (N2→NH3)

Inorganic synthesis for Biomimetic modeling of FeMoco

Nitrogen fixation — the conversion of atmospheric dinitrogen (N2) to ammonia (NH3) — is one of the most chemically challenging transformations in nature, requiring cleavage of the N≡N triple bond (bond dissociation energy ~225 kcal/mol). Breaking the first bond alone requires ~98 kcal/mol. The industrial Haber–Bosch process accomplishes this under extreme conditions, using hydrogen gas generated from the Water-Gas Shift and Methane Steam Reforming processes. All of these industrial processes, however, require enormous energy resources to maintain the high temperatures and pressures, consuming roughly 1–2% of the global energy supply. In contrast, the nitrogenase enzymes achieve the same transformation at ambient temperature and pressure utilizing aqueous protons (H+) and electrons in lieu of hydrogen gas.

At the heart of nitrogenase activity is a double-cuboidal iron-sulfur cluster cofactor. My graduate research focused on synthetic modeling of the active site of Molybdenum-dependent nitrogenase, called FeMoco, with the goal of understanding how its unique architecture — including an interstitial carbide and variable metal-sulfur environments — facilitate the nitrogen fixation with protons and electrons. By developing new synthetic pathways to incorporate sulfide, carbide, and heterometals into iron clusters, I generated a family of biomimetic complexes that emulate key structural and electronic features of FeMoco. These efforts have provided design strategies for synthetic construction of structural models of nitrogenase which continue to be used today, and may one day offer potential routes to functional alternatives to Haber–Bosch catalysis.

Old dog, new tricks : repurposing iron-carbide-carbonyl clusters as precursors for structural modeling of the nitrogenase cofactor, The University of Texas at Austin, 2020. DOI: 10.26153/tsw/13865

Active site cluster of Mo-dependent nitrogenase (FeMoco) with key structural and electronic characteristics indicated.

Synthetic Conversions of Cluster Core

A key objective of this project was to develop synthetic precursors that emulate the FeMoco core structure — in particular, its highly unique 6-coordinate interstitial carbide atom. Using a modular family of iron-carbide-carbonyl clusters originally reported in the 1960s, a series of interconversion pathways were developed to manipulate the cluster core and facilitate changes in carbide coordination or cluster oxidation state. This provided pathways to forming 6-, 5-, and 4-coordinate carbide motifs, which could serve as precursors for heterometal incorporation. Through iterative multistep syntheses, we successfully incorporated Mo into a [Fe5Mo(μ6-C)] cluster core, thus establishing synthetic access to cluster architectures with tunable metal composition. The Mo-containing cluster did demonstrate catalytic competence for diphenylacetylene activation via a hydride intermediate, providing some preliminary functional analogy to FeMoco, in addition to the structural biomimetics.

Publications:

Inorg. Chem. 2018, 57, 20–23. DOI: 10.1021/acs.inorgchem.7b02615
Inorg. Chem. 2017, 56, 5998–6012. DOI: 10.1021/acs.inorgchem.7b00741

The Electronic Role of the Interstitial Carbide: Evidence from X-ray Spectroscopy

To better understand how electron density is distributed across the cluster and to evaluate the role of the interstitial carbide in FeMoco analogues, we conducted a detailed X-ray spectroscopic analysis on the clusters developed as described above, specifically the homonuclear Fe6 and Fe5 clusters and a heteronuclear Fe5Mo cluster. Using a combination of X-ray Absorption Spectroscopy (XAS), Mössbauer spectroscopy, and Valence-to-Core X-ray Emission Spectroscopy (VtC-XES), we showed that the carbide ligand plays a central role in charge delocalization, enhancing electron density at iron sites despite overall cluster oxidation. Notably, Mössbauer spectra revealed that the formally more oxidized Fe5 cluster actually exhibited lower iron oxidation states compared to Fe6, highlighting the importance of metal–carbide covalency in stabilizing redox-active states. Spectroscopic data, paired with DFT calculations, provided a nuanced view of metal–metal and metal–carbide bonding interactions and clarified how structural changes (e.g., Mo substitution, ligand coordination) influence electronic structure. These findings offer compelling insight into how the interstitial carbide in FeMoco may function as an electronic buffer, helping the biological cluster maintain redox flexibility across multiple electron-transfer events.

Inorg. Chem. 2019, 58, 12918–12932. DOI: 10.1021/acs.inorgchem.9b01870

Electrophilic Sulfur Insertion into Fe–C Cores

A major challenge in synthetic modeling of the nitrogenase FeMoco is the simultaneous incorporation of both an interstitial carbide and inorganic sulfides — a structural hallmark of the biological cofactor. In this project, several approaches were taken in pursuit of this motif. Finally, we developed a synthetic strategy to achieve single-step sulfur insertion into iron-carbide-carbonyl clusters using electrophilic sulfur sources such as S₈ and S₂Cl₂. This method yielded a series of previously inaccessible iron-sulfide-carbide clusters, including sulfide-bridged dimers and mononuclear thiolate-substituted analogues. One key advance was the formation of a sulfocarbide motif ({C–S}⁴⁻), which models a proposed biosynthetic intermediate in FeMoco maturation via the NifB protein. These clusters represent the first synthetic systems that simultaneously incorporate all three essential features of FeMoco: iron, sulfide, and a central carbide. The work also demonstrated that sulfurization promotes higher oxidation states at Fe centers, modulating both the electronic structure and reactivity of the cluster. This approach opens new pathways for stepwise construction of FeMoco-inspired architectures and provides valuable insights into how sulfur and carbon ligands cooperate to stabilize redox-active, multimetallic cores.

Publications:

Catalysts 2020, 10, 1317. DOI: 10.3390/catal10111317
Angew. Chem. Int. Ed. 2021, 60, 20321–20327. DOI: 10.1002/anie.202011517

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