Residue-Orientation-Dependent Dynamics and Selectivity of Active Pocket in Microbe Class I Terpene Cyclases

Abstract

Microbe class I terpene cyclases (TPCs) are responsible for deriving numerous functionally and structurally diverse groups of terpenoid natural products. The conformational change of their active pockets from "open" state to "closed" state upon substrate binding has been clarified. However, the key structural basis relevant to this active pocket dynamics and its detailed molecular mechanism are still unclear. In this work, on the basis of the molecular dynamics (MD) on two microbe class I TPCs (SdS and bCinS), we propose that the active pocket dynamics is highly dependent on the residue orientation of two conserved structural bases R-D dyad and X-R-D triad, rather than the previously suggested flexibility of kink region. Actually, we considered that the flexibility of kink region is synchronous with the R residue orientation of the X-R-D triad, which could regulate the entrance size of active pocket and thus affect the substrate selectivity of active pocket by utilizing the promiscuity of the X-R-D triad. Furthermore, to better understand the function of the two structural bases, two intelligible models of "PPi catcher-locker" and "selector-PPi sensor-orienter" are proposed to, respectively, describe the R-D dyad and X-R-D triad and broadened to more microbe class I TPCs. These findings exhibit the dynamics of active pocket inaccessible in static crystal structures and provide useful structural basis knowledge for further design of microbe class I TPCs with different cyclization ability.

Insights into the enzymatic catalytic mechanism of bCinS: the importance of protein conformational change

“…Overall, combining with our previous study of substrate binding state, 20 the whole chemical catalysis process of bCinS could be concluded in Fig. 11.…”

“…17,18 In the case of Streptomyces clavuligerus 1,8-cineole synthase (bCinS, a typical microbe class I TPC), 19 the crystal conformation with the binding of two Mg 2+ ions (2Mg 2+ ) and the substrate analogue 2-fluoroneryl pyrophosphate (FNPP) can be regarded as the "half-closed" state (Fig. 1a), 19,20 and the state with 3Mg 2+ and FNPP is "closed" where the residue R174 forms a salt-bridge interaction with a diphosphate moiety (PPi) to further close the active site pocket (Fig. 1b).…”

“…Many experimental and theoretical studies have demonstrated the contribution of cofactors, protonation states, and conserved Arg residues (R174 in bCinS) to the PPi cleavage. 1,16,21 In previous classical molecular dynamics (MD) simulations, we have revealed the conformational change of bCinS from the "open" state to the "closed" state, which mainly focused on the process from the substrate entering to the closure of the active pocket, 20 but the key regulators related to its chemical catalysis process remain unclear. Recent studies supposed that the carbonyl in the kink region defined as the "effector" (G178-O in bCinS) can trigger the PPi cleavage by interacting with the π* molecular orbital of the C2-C3 double-bond (Fig.…”

Mono-phosphorylation at Ser4 of barrier-to-autointegration factor (Banf1) significantly reduces its DNA binding capability by inducing critical changes in its local conformation and DNA binding surface

“…Similar molecular modelling techniques have been applied to successfully unravel the molecular-and/or atomic-level mechanisms of biological systems that were challenging for then-current experimental techniques, [19][20][21][22][23][24] including designing engineered transmembrane proteins recently reported by us, 25 binding of transcription factors to DNA, 26 complicated dynamics of enzymes, 27,28 importance of WPD-Loop sequence for activity and structure in protein tyrosine phosphatases, 29 the surface adhesion mechanisms of COVID-19, 30 the nanomechanical features of coronavirus spike proteins, 31,32 and the conformational dynamics of proteins related to diseases. [33][34][35][36][37][38][39] Computational models and methodology…”