Yong Zhang

Mössbauer spectroscopy is among the best techniques for probing molecular structures of iron complexes and proteins. Extracting structural insights from experimental spectra often necessitates accurate DFT predictions of isomer shift (IS) and quadrupole splitting (QS) parameters. However, substantial computational cost and sensitivity to the choice of functional and basis set limit their utility for high-throughput structural screening. To address this gap, various traditional machine learning (ML) and deep learning (DL) models were investigated, using a curated experimental data set of 522 diverse compounds. After engineering models and descriptors, the DL model Chemprop has the highest coefficient of determination (R2) for the test data set (0.92 for IS; 0.73 for QS), with R2 = 0.98/0.91 for IS/QS in the overall data set. Validation of this model corroborates its high, close-to-DFT accuracy for IS predictions, and also shows a significant improvement room for QS predictions. Its predictions of several iron sites in both heme and nonheme proteins and both first- and second-coordination sphere models with challenging and/or unique novel structural features replicated previous DFT structural predictions that are in excellent agreement with experiment, which is a significant application given that no protein data was used to train the ML model. This approach achieved a 218-fold reduction in computational time compared to DFT calculations with same geometries. Results from this first ML work on Mössbauer data of molecular iron complexes will facilitate rapid structural screening previously impractical for large sets of iron complexes and iron-protein binding site structures.

Nitric oxide (NO) insertion into a metalloporphyrin rhodium–carbon bond is observed when crystals of (TPP)Rh(C₆H₄Cl)(CH₂Cl₂)_x (TPP = tetraphenylporphyrinato dianion) are exposed to NO gas over a two week period. The NO-inserted product was identified by X-ray crystallography as (TPP)Rh(ONC₆H₄Cl-o)(CH₂Cl₂)_0.5·(TPP)Rh(C₆H₄Cl-p/m)(CH₂Cl₂)_0.5, in which NO inserted selectively into the ortho-Cl aryl isomer but not into the para-/meta-Cl aryl isomers. Density functional theory calculations predict a higher reactivity for the ortho-Cl isomer and thus provide a rationale for the preferred NO insertion into the potentially more reactive ortho-Cl isomer under our reaction conditions. Whereas NO insertion reactions are well-known for coordination–organometallic compounds, this result represents the first such NO insertion into a metalloporphyrin metal–carbon bond for any metal.

Nitroxyl (HNO) is a biologically active nitrogen oxide with therapeutic potential for several disorders, but its endogenous formation remains poorly understood. Previous experimental work showed that hydroxylamine (NH₂OH), a possible physiological precursor, was oxidized by hydrogen peroxide (H₂O₂) activated heme proteins to produce HNO. In this study, we employed both density functional theory (DFT) and hybrid quantum mechanics and molecular mechanics (QM/MM) calculations to investigate the reaction mechanisms of HNO generation from NH₂OH via two representative heme proteins, myoglobin (Mb) and catalase (CAT), with neutral and negative axial ligands. Various structural models to successively include more components of the proteins were studied to provide more detailed insights into the protein environment effect. Our results reveal a stepwise double hydrogen atom abstraction pathway initiated by the ferryl compound I (Cpd I) species after the H₂O₂ treatment, with the first H-abstraction as the rate-determining step. Mb, with a neutral histidine axial ligand, exhibited a reaction barrier lower than that of CAT (which contains a negatively charged tyrosine ligand), which was in good agreement with the observed experimental reactivity trend. Protein environment effects, including distal hydrogen bonding and steric hindrance, were found to modulate both proton affinity and electron transfer, influencing the reaction barriers. QM/MM calculations confirmed that while key residues near the heme center can enhance HNO formation through hydrogen bonding, spatial constraints in the full protein environment can raise the reaction barrier. These mechanistic insights explain experimentally observed differences in HNO yields and highlight how protein structure and heme coordination influence nitroxyl biosynthesis.

Recently, engineered heme proteins have been developed as excellent catalysts for carbene transfer reactions to facilitate sustainable chemical synthesis. Computational mechanistic investigations provide significant insights into the understanding of various aspects of these reactions. Both density functional theory (DFT) calculations with accurate predictions of many experimental spectroscopic properties and high-level computational studies were employed to elucidate the electronic structures of key heme carbene reaction intermediates. Effects of heme catalysis, coordination modes, and structural components on heme carbene formation via diazo and non-diazo precursors were revealed. Besides the basic mechanisms discovered for multiple heme carbene transfer reactions such as cyclopropanations and X–H insertions (X = Si, C, N) with many experimental features reproduced, details of how the carbene substituent, axial ligand, porphyrin substituent, and substrate substituent influence the reactivity were also examined to help future reaction design. In addition, the mechanistic origins of stereoselectivities and chemoselectivities for some important experimental heme carbene transfer reactions were disclosed to further advance this field. The interesting mechanistic results summarized in this review will facilitate the application of new-to-nature heme based biocatalysis.

A recent experimental study by Arnold’s research group (Nature 2019, 565, 67–72) showed the engineered cytochrome P450 enzyme (cytochrome P411), with acceptor-only carbenes, offers a new sustainable alternative to metal catalysts that have traditionally dominated C–H functionalization. However, there is still no detailed mechanistic understanding thus far. In this study, a series of DFT calculations were performed to uncover the basic reaction mechanism as well as insights into the observed carbene substituent effect behind this novel reaction. Computational results showed that the reaction follows the radical hydrogen atom transfer (HAT) mechanism, which is consistent with experimental work. The electron-withdrawing carbene substituent together with the negatively charged axial ligand plays an important role in steering the reaction toward this mechanism. The more favorable experimental reactivity with the carbene derived from ethyl diazoacetate than diazoacetone was also reproduced. It was found to originate from the additional hydrogen bonds with the carboxylate substituent of the carbene, which further stabilizes the transition state toward a more facile HAT reaction. This quantum chemical study provides useful molecular-level insights that help shed light on the future design of the biocatalysts to contribute to the development of efficient, sustainable C–H functionalization approaches.

HNO plays an important role in many biological processes related to vasodilation, regulation of enzyme activities, and neurological functions. The understanding of enzymatic oxidative HNO-releasing pathways in biology remains scarce. We investigated HNO formation from a well-established small-molecule drug hydroxyurea catalyzed by horseradish peroxidase (HRP). Density functional theory results reveal two sequential proton-coupled electron transfers from hydroxyurea to HRP Compound I as the most favorable mechanism for HNO generation, which was found to be kinetically feasible and thermodynamically favorable. This is consistent with its experimentally observed reactivity. Moreover, the large active site model study uncovered interesting conformation changes involving HRP’s amino acid H-bonding network through the reaction pathway, which were employed to anchor the evolving substrate via its key CONH₂ moiety. Detailed computational analysis reveals some useful structural properties for future drug development. In addition, this study presents the first computational investigation of the full reaction mechanism of acyl nitroso hydrolysis for HNO generation, supporting the experimentally observed facile reactivity. Overall, this study reveals, for the first time, the detailed computational reaction mechanism of enzymatic oxidative and hydrolytic HNO generation from a clinical drug, providing structural features beneficial for future drug design.

Nitric oxide (NO) is an important molecule that regulates many physiological processes in humans and plants and contributes to the formation of greenhouse gases. Bacterial NO reductases utilize a di-Fe heme/nonheme active site to couple two NOs to generate nitrous oxide (N₂O) via a two-electron mechanism. Here, we report a previously unexplored Cr porphyrin NO complex with a Lewis acid (LA) BF₃ for the NO reduction reaction. Density functional theory calculations were first employed to reveal its reaction mechanism with a reasonable barrier for experimental realization. Subsequent experimental synthesis work confirms this reactivity and reports the first nitrosyl Cr porphyrin X-ray crystal structure. Theoretical analysis uncovered a distinctive reaction feature for the Cr system compared to Fe and Co porphyrins: the electron transfer from the metal to the bound NO occurs before LA binding. A comparative study of the NO coupling mechanisms with the three representative metals suggests that the metal reduction potential should be finely tuned, as found in previous studies of NOR enzymatic systems. Overall, this study offers new theoretical and experimental insights to further facilitate the development of alternative NO reduction compounds with biological, environmental, and industrial applications.

Calculations provide the first comprehensive mechanistic study of carbene N–H insertion for His-ligated heme biocatalysts, as validated via new experiments.

Engineered heme proteins exhibit excellent sustainable catalytic carbene transfer reactivities toward olefins for value-added cyclopropanes. However, unactivated and electron-deficient olefins remain challenging in such reactions. To help design efficient heme-inspired biocatalysts for these difficult situations, a systematic quantum chemical mechanistic study was performed to investigate effects of olefin substituents, non-native amino acid axial ligands, and natural and non-natural macrocycles with the widely used ethyl diazoacetate. Results show that electron-deficient substrate ethyl acrylate has a much higher barrier than the electron-rich styrene. For styrene, the predicted barrier trend is consistent with experimentally used heme analogue cofactors, which can significantly reduce barriers. For ethyl acrylate, while the best non-native axial ligand only marginally improves the reactivity versus the native histidine model, a couple of computationally studied macrocycles can dramatically reduce barriers to the level comparable to styrene. These results will facilitate the development of better biocatalysts in this area.

Some pathogens use heme‐containing nitric oxide reductases (NORs) to reduce NO to N₂O as their defense mechanism to detoxify NO and reduce nitrosative stress. This reduction is also significant in the global N cycle. Our previous experimental work showed that Fe and Co porphyrin NO complexes can couple with external NO to form N₂O when activated by the Lewis acid BF₃. A key difference from conventional two‐electron enzymatic reaction is that one electron is sufficient. However, a complete understanding of the entire reaction pathways and the more favorable reactivity for Fe remains unknown. Here, we present a quantum chemical study to provide such information. Our results confirmed Fe's higher experimental reactivity, showing advantages in all steps of the reaction pathway: easier metal oxidation for NO reduction and N−O cleavage as well as a larger size to expedite the N/O coordination mode transition. The Co system, with a similar product energy as the enzyme, shows potential for further development in catalytic NO coupling. This work also offers the first evidence that this new one‐electron NO reduction is both kinetically competitive and thermodynamically more favorable than the native pathway, supporting future initiatives in optimizing NO reduction agents in biology, environment, and industry.