Substrate Positioning Dynamics Involves a Non-Electrostatic Component to Mediate Catalysis

Jiang, Yaoyukun; Ding, Ning; Shao, Qianzhen; Stull, Sebastian L.; Cheng, Zihao; Yang, Zhongyue J.

doi:10.1021/acs.jpclett.3c02444

Cited by 4 publications

(2 citation statements)

References 93 publications

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“…For enzyme HiC, the charges of atoms PET@C1 and PET@O2 are similar in both Re -face and Si -face binding modes, while the intrinsic electric field along the PET@O2-PET@C1 axis in Re -face binding (−2.4 MV cm –1 ) is weaker than that in Si -face binding (−9.3 MV cm –1 ). We note that the interplay between the intrinsic electric field and substrate positioning dynamics that mediate the depolymerization efficiency of PETase could be explored in the near future . Nevertheless, the present results evidenced the contribution of the intrinsic electric field on the lower energy barriers of HiC’s Re -face binding in step ii compared to that of FAST-PETase and LCC ICCG .…”

Section: Resultssupporting

confidence: 77%

Prochiral Selectivity in Enzymatic Polyethylene Terephthalate Depolymerization Revealed by Computational Modeling

Zheng,

Zhu,

et al. 2024

ACS EST Eng.

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Enzyme catalysis has shown its great power in dealing with global poly(ethylene terephthalate) (PET) waste. However, it is still challenging to design a super enzyme that can treat the sheer volume of worldwide PET waste. Without a complete understanding of the catalytic mechanism, it will be difficult to reach this important goal. Here, we systematically study the PET depolymerization mechanism catalyzed by structurally different hydrolases. The role of fleeting chiral intermediates was proved to be crucial. We observed different prochiral selectivities among these PET hydrolases. While most hydrolases favor Si-face binding, a few hydrolases (e.g., Humicola insolens cutinase) mainly adapt Re-face binding. Interestingly, we found that Si-face binding leads to higher activity than Re-face binding in all of the studied hydrolases. This Si-face selectivity originates from the difficulty of proton transfer from catalytic histidine residue to the substrate and the less stability of the oxyanion hole. Since the Si-face binding ratio ranges from 0 to 95%, we infer that all these hydrolases are not perfectly evolved to degrade PET. Our in silico results highlight that enlarging binding site residues (e.g., Leu66 and Asn69) will enhance enzymatic depolymerization, which was further confirmed by our in vitro experiments where both Leu66Phe and Asn69Phe show significantly increased PET hydrolysis activity. Hopefully, this work will aid the future rational design of super enzymes to fight PET pollution.

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Section: Resultssupporting

confidence: 77%

Prochiral Selectivity in Enzymatic Polyethylene Terephthalate Depolymerization Revealed by Computational Modeling

Zheng,

Zhu,

et al. 2024

ACS EST Eng.

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“…Despite the aforementioned mechanistic insights, the molecular mechanism of the P450-catalyzed biosynthesis of DKPs is not fully elucidated, especially regarding the protein environment effects in dictating the regio- and stereoselective C–N and C–C coupling. Extensive studies have shown that the protein environment is key to the activity and selectivity of enzyme catalysis, while the neglect of such an effect in the QM model calculations may give biased descriptions of the kinetic and thermodynamic properties of enzymatic processes. ,− In this study, we reexamined the molecular mechanism of the P450 NascB with the combined MD simulations , and quantum mechanical/molecular mechanical (QM/MM) calculations. − Particularly, we systematically explored all conceivable catalytic pathways originating from two distinct binding modes of Substrate 1. Our multiscale calculations reveal that pathway B, which involves a conformational movement of the substrate radical, is the most favorable reaction pathway.…”

Section: Introductionmentioning

confidence: 99%

Substrate Conformational Switch Enables the Stereoselective Dimerization in P450 NascB: Insights from Molecular Dynamics Simulations and Quantum Mechanical/Molecular Mechanical Calculations

Zhou,

Feng,

Wang

et al. 2024

JACS Au

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P450 NascB catalyzes the coupling of cyclo-(l-tryptophan-l-proline) (1) to generate (−)-naseseazine C (2) through intramolecular C–N bond formation and intermolecular C–C coupling. A thorough understanding of its catalytic mechanism is crucial for the engineering or design of P450-catalyzed C–N dimerization reactions. By employing MD simulations, QM/MM calculations, and enhanced sampling, we assessed various mechanisms from recent works. Our study demonstrates that the most favorable pathway entails the transfer of a hydrogen atom from N7–H to Cpd I. Subsequently, there is a conformational change in the substrate radical, shifting it from the Re-face to the Si-face of N7 in Substrate 1. The Si-face conformation of Substrate 1 is stabilized by the protein environment and the π–π stacking interaction between the indole ring and heme porphyrin. The subsequent intermolecular C3–C6′ bond formation between Substrate 1 radical and Substrate 2 occurs via a radical attack mechanism. The conformational switch of the Substrate 1 radical not only lowers the barrier of the intermolecular C3–C6′ bond formation but also yields the correct stereoselectivity observed in experiments. In addition, we evaluated the reactivity of the ferric-superoxide species, showing it is not reactive enough to initiate the hydrogen atom abstraction from the indole NH group of the substrate. Our simulation provides a comprehensive mechanistic insight into how the P450 enzyme precisely controls both the intramolecular C–N cyclization and intermolecular C–C coupling. The current findings align with the available experimental data, emphasizing the pivotal role of substrate dynamics in governing P450 catalysis.

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Unlocking the potential of enzyme engineering via rational computational design strategies

Zhou,

Tao,

Shen

et al. 2024

Biotechnology Advances

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Substrate Positioning Dynamics Involves a Non-Electrostatic Component to Mediate Catalysis

Cited by 4 publications

References 93 publications

Prochiral Selectivity in Enzymatic Polyethylene Terephthalate Depolymerization Revealed by Computational Modeling

Prochiral Selectivity in Enzymatic Polyethylene Terephthalate Depolymerization Revealed by Computational Modeling

Substrate Conformational Switch Enables the Stereoselective Dimerization in P450 NascB: Insights from Molecular Dynamics Simulations and Quantum Mechanical/Molecular Mechanical Calculations

Unlocking the potential of enzyme engineering via rational computational design strategies

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