@article{gering_li_tang_swartz_chang_makris_2023, title={A Ferric-Superoxide Intermediate Initiates P450-Catalyzed Cyclic Dipeptide Dimerization}, volume={8}, ISSN={["1520-5126"]}, url={https://doi.org/10.1021/jacs.3c04542}, DOI={10.1021/jacs.3c04542}, abstractNote={The cytochrome P450 (CYP) AspB is involved in the biosynthesis of the diketopiperazine (DKP) aspergilazine A. Tryptophan-linked dimeric DKP alkaloids are a large family of natural products that are found in numerous species and exhibit broad and often potent bioactivity. The proposed mechanisms for C-N bond formation by AspB, and similar C-C bond formations by related CYPs, have invoked the use of a ferryl-intermediate as an oxidant to promote substrate dimerization. Here, the parallel application of steady-state and transient kinetic approaches reveals a very different mechanism that involves a ferric-superoxide species as a primary oxidant to initiate DKP-assembly. Single turnover kinetic isotope effects and a substrate analog suggest the probable nature and site for abstraction. The direct observation of CYP-superoxide reactivity rationalizes the atypical outcome of AspB and reveals a new reaction manifold in heme enzymes.}, journal={JOURNAL OF THE AMERICAN CHEMICAL SOCIETY}, author={Gering, Hannah E. and Li, Xiaojun and Tang, Haoyu and Swartz, Paul D. and Chang, Wei-Chen and Makris, Thomas M.}, year={2023}, month={Aug} }
 @article{ushimaru_cha_shimo_li_paris_mori_miyamoto_coffer_uchiyama_guo_et al._2023, title={Mechanistic Analysis of Stereodivergent Nitroalkane Cyclopropanation Catalyzed by Nonheme Iron Enzymes}, volume={145}, ISSN={["1520-5126"]}, DOI={10.1021/jacs.3c08413}, abstractNote={BelL and HrmJ are α-ketoglutarate-dependent nonheme iron enzymes that catalyze the oxidative cyclization of 6-nitronorleucine, resulting in the formation of two diastereomeric 3-(2-nitrocyclopropyl)alanine (Ncpa) products containing trans-cyclopropane rings with (1'R,2'R) and (1'S,2'S) configurations, respectively. Herein, we investigate the catalytic mechanism and stereodivergency of the cyclopropanases. The results suggest that the nitroalkane moiety of the substrate is first deprotonated to produce the nitronate form. Spectroscopic analyses and biochemical assays with substrates and analogues indicate that an iron(IV)-oxo species abstracts proS-H from C4 to initiate intramolecular C-C bond formation. A hydroxylation intermediate is unlikely to be involved in the cyclopropanation reaction. Additionally, a genome mining approach is employed to discover new homologues that perform the cyclopropanation of 6-nitronorleucine to generate cis-configured Ncpa products with (1'R,2'S) or (1'S,2'R) stereochemistries. Sequence and structure comparisons of these cyclopropanases enable us to determine the amino acid residues critical for controlling the stereoselectivity of cyclopropanation.}, number={44}, journal={JOURNAL OF THE AMERICAN CHEMICAL SOCIETY}, author={Ushimaru, Richiro and Cha, Lide and Shimo, Shotaro and Li, Xiaojun and Paris, Jared C. and Mori, Takahiro and Miyamoto, Kazunori and Coffer, Lindsay and Uchiyama, Masanobu and Guo, Yisong and et al.}, year={2023}, month={Oct}, pages={24210–24217} }
 @article{li_xue_guo_chang_2022, title={Mechanism of Methyldehydrofosmidomycin Maturation: Use Olefination to Enable Chain Elongation}, volume={144}, ISSN={["1520-5126"]}, DOI={10.1021/jacs.2c01924}, abstractNote={Utilization of mononuclear iron- and 2-oxoglutarate-dependent (Fe/2OG) enzymes to enable C-H bond functionalization is a widely used strategy to diversify the structural complexity of natural products. Besides those well-studied reactions including hydroxylation, epoxidation, and halogenation, in the biosynthetic pathway of dehydrofosmidomycin, an Fe/2OG enzyme is reported to catalyze desaturation, alkyl chain elongation, along with demethylation in which trimethyl-2-aminoethylphosphonate is converted into methyldehydrofosmidomycin. How this transformation takes place is largely unknown. Herein, we characterized the reactive species, revealed the structure of the reaction intermediate, and used mechanistic probes to investigate the reaction pathway and mechanism. These results led to the elucidation of a two-step process in which the first reaction employs a long-lived Fe(IV)-oxo species to trigger C═C bond installation. During the second reaction, the olefin installed in situ enables C-C bond formation that is accompanied with a C-N bond cleavage and hydroxylation to furnish the alkyl chain elongation and demethylation. This work expands the reaction repertoire of Fe/2OG enzymes by introducing a new pathway to the known C-C bond formation mechanisms utilized by metalloenzymes.}, number={18}, journal={JOURNAL OF THE AMERICAN CHEMICAL SOCIETY}, author={Li, Xiaojun and Xue, Shan and Guo, Yisong and Chang, Wei-Chen}, year={2022}, month={May}, pages={8257–8266} }
 @article{li_shimaya_dairi_chang_ogasawara_2021, title={Identification of Cyclopropane Formation in the Biosyntheses of Hormaomycins and Belactosins: Sequential Nitration and Cyclopropanation by Metalloenzymes}, volume={12}, ISSN={["1521-3773"]}, DOI={10.1002/anie.202113189}, abstractNote={Hormaomycins and belactosins are peptide natural products that contain unusual cyclopropane moieties. Bioinformatics analysis of the corresponding biosynthetic gene clusters showed that two conserved genes, hrmI / belK and hrmJ / belL , were potential candidates for catalyzing cyclopropanation. Using in vivo and in vitro assays, the functions of HrmI/BelK and HrmJ/BelL were established. HrmI and BelK, which are heme oxygenase-like dinuclear iron enzymes, catalyze oxidation of the ε-amino group of L-lysine to afford L-6-nitronorleucine. Subsequently, HrmJ and BelL, which are iron- and α-ketoglutarate-dependent oxygenases, effectively convert L-6-nitronorleucine to 3-( trans -2-nitrocyclopropyl)alanine through C4-C6 bond installation. These observations disclose a novel pathway of cyclopropane ring construction and exemplify the new chemistry involving metalloenzymes in natural product biosynthesis.}, journal={ANGEWANDTE CHEMIE-INTERNATIONAL EDITION}, author={Li, Xiaojun and Shimaya, Ryo and Dairi, Tohru and Chang, Wei-chen and Ogasawara, Yasushi}, year={2021}, month={Dec} }