Enzyme Promiscuity versus Fidelity in Two Sesquiterpene Cyclases (TEAS versus ATAS)

Abstract

Catalytic promiscuity, as a modern and imperfect understanding concept in enzyme catalysis community, is prevailing in plant-derived sesquiterpene cyclases (FPPC) and highly related to the chemical diversity of sesquiterpenoid natural products. Both the Nicotiana tabacum 5-epi-aristolochene synthase (TEAS) and Aspergillus terreus aristolochene synthase (ATAS) belong to FPPC, involve the same reaction pathway, and yield their ultimate main product (aristolochene) with different stereochemistries. The catalytic promiscuity of TEAS and fidelity of ATAS have been observed in previous experimental studies, but the detailed catalytic mechanism is still not clear. Herein, by employing the quantum classical multiscale molecular dynamics simulations and site-directed mutagenesis experiments, the complete enzyme catalytic pathways from the substrate to aristolochene and various side products (in TEAS) are investigated and the important mechanism insights are included: (1) the PPi moiety (diphosphate group, released from the substrate) would further act as the general acid/base in both TEAS and ATAS enzyme catalysis, which likely plays a general role in FPPC. (2) The Asp444−Tyr520 dyad acts as an additional general acid/base residue pair to increase promiscuity in TEAS. (3) The enriched aromatic residues are essential for the catalytic fidelity of ATAS. Finally, we further discuss the three critical chemical control factors which are proposed to be responsible for the catalytic promiscuity and fidelity in most FPPC, that is, substrate folding mode, intermediate flexibility, and key residue, owing to the more or less plasticity of the active pocket in various FPPC.

Molecular Basis for Sesterterpene Diversity Produced by Plant Terpene Synthases

“…Which residues are involved in the final deprotonation reaction? Computational-guided methods (e.g., quantum mechanical/molecular mechanical–molecular dynamic simulations), together with the structural features of AtTPS18, will help address these interesting questions ( Zhang et al., 2020 ).…”

Structural studies reveal flexible roof of active site responsible for ω-transaminase CrmG overcoming by-product inhibition

“…All the QM/MM simulations were run with QChem4.0 40 and Tinker 41 programs. The whole protocol is transplanted from our previous studies and has been carefully validated [42][43][44] . Additional a The values in parentheses refer to statistics in the highest bin.…”

“…All the QM/MM simulations were run with QChem4.0 40 and Tinker 41 programs. The whole protocol is transplanted from our previous studies and has been carefully validated 42 – 44 . Additional benchmark tests on the computational level (functionals and basis sets), choice of QM region and simulation time are described in Supporting Information (Supplementary Figs.…”

Predictive Engineering of Class I Terpene Synthases Using Experimental and Computational Approaches

“…Deprotonation is the most common termination mechanism, but the catalytic base for proton abstraction is not always obvious. Computational approaches supplied further evidence for the involvement of PP, [76] the employment of non‐traditional bases such as Ser, [77] as well as the participation of multiple bases in the final deprotonation step [59] . The use of multiple acid/base pairs has been implicated in increased promiscuity in TSs, offering a multitude of termination options [76] .…”