Guojun Zheng

Chiral C₃-aminomethyl cyclopentanones are key intermediates in organic chemistry, yet their synthesis remains challenging. In this study, we present a kinetic resolution catalyzed by ene-reductases─OYE1 mutants─enabling C₃-aminomethyl cyclopentanones to be resolved into R-C₃-aminomethyl cyclopentanones (up to 99% e.e.) and corresponding α,β-unsaturated cyclopentanones via desaturation reactions. Remarkably, the reaction remains viable at the gram scale even with lower catalyst loadings, demonstrating potential for industrial-scale implementation.

A comprehensive understanding of the biophysical parameters that dictate high catalytic efficiency in enzymes is essential for advancing both fundamental enzymology and its applications. Experimental evidence suggests that protein dynamics play a pivotal role in transiently shaping active site configurations, facilitating the efficient traversal of reaction barriers. In a previous study, protein engineering led to the development of a triple mutant of myoglobin, which enabled the successful synthesis of an array of chiral flavanone compounds through myoglobin‐mediated carbene transfer. To gain deeper insights into the molecular mechanisms underlying the evolution of structural dynamics that contribute to the accelerated catalytic properties, we first performed hydrogen‐deuterium exchange mass spectrometry (HDX‐MS) analyses on both the wild‐type myoglobin and the engineered triple mutant in the presence and absence of a substrate analog to elucidate conformational changes and impacts of mutations on protein flexibility and functional dynamics. HDX‐MS analysis identified distinct regions of the protein, both proximal and distal to the mutation sites, which exhibited differential HDX behaviors in response to either ligand binding or mutation, thereby providing insights into the structural and dynamic evolution of the mutant. We postulate that the mutation reconfigures the conformational ensemble of the protein, thereby promoting favorable conformational sampling and enhancing the efficiency of the catalyzed reaction. Computational studies further support this conclusion, providing additional insights into the structural and dynamic factors influencing enzymatic efficiency. This study highlights the critical role of protein structural dynamics in evolved enzymes, underscoring the potential of probing and harnessing these dynamics for advancements in protein engineering and redesign.

Engineered myoglobin variants were able to activate C(sp³)–H under mild conditions via mediated nitrene transfer and intramolecular cyclisation to form highly enantioselective 2-oxazolidinone and γ-lactams.

In organic chemistry, the umpolung reaction is an important concept that significantly expands the chemical space and provides unique structures. Although prior research focused on constructing complex molecular frameworks using phase transfer catalysts, the umpolung reaction is well known for its roles in 1,4-conjugate addition and Stetter reactions. Herein, we reported a novel enzymatic approach using immobilized transaminase CV2025 to catalyze the umpolung conjugate addition reaction. This method facilitates the nucleophilic addition of 2-(nitrophenyl)-N-(trifluoromethyl) imine to cyclopentanone, efficiently synthesized a complex imine and its derivatives up to 78% yield. Unlike traditional methods, this enzymatic strategy eliminates the need of a transition metal or chemical (acid, base) catalyst, broadens the substrate scope, and operates under milder conditions. Mechanistic studies, supported by molecular dynamics calculations and alanine scanning mutagenesis, identified key residues influencing regioselectivity. This study presents a sustainable, enzyme-catalyzed method for constructing complex frameworks with broad potential in pharmaceutical synthesis and beyond.

Fucosyltransferase 2 (FUT2) gene has been extensively reported to play its role in potential gut microbiota changes and norovirus susceptibility. The normal activity of FUT2 has been found to be disrupted by non-synonymous single nucleotide polymorphisms (nsSNPs) in its gene. To explore the possible mutational changes and their deleterious effects, we employed state-of-the-art computational strategies. Firstly, nine widely-used bioinformatics tools were utilized for initial screening of possibly deleterious nsSNPs. Subsequently, the structural and functional effects of screened nsSNPs on FUT2 were evaluated by utilizing relevant computational tools. Following this, the two shortlisted nsSNPs, including G149S (rs200543547) and V196G (rs367923363), were further validated by their molecular docking with norovirus capsid protein, VP1. As compared to wild-type, the higher stability and lower binding energy scores of the both the mutants indicated their stable binding with VP1, which ultimately leads to norovirus implications. These docking results were further verified by a comprehensive computational approach, molecular dynamic simulation, which gave results in the form of lower RMSD, RMSF, RoG, and hydrogen bond values of both the mutants, depicted in relevant graphs. Overall, this research explores and validated the two FUT2 nsSNPs (G146S and V196G), which may possibly linked with the norovirus susceptibility and gut microbiota changes. Moreover, our findings highlights the value of computational strategies in mutational analysis and welcomes any further experimental validation.