Linalool dehydratase/isomerase (LinD) from Castellaniella defragrans is a bifunctional enzyme that catalyzes the hydration of β-myrcene to (S)-linalool and the isomerization of (S)-linalool to geraniol. In this paper, on the basis of recently obtained crystal structures, the catalytic mechanism of LinD has been explored by a combined quantum mechanics and molecular mechanics (QM/MM) approach. Two computational models have been constructed. Model I (LinD-linalool) was derived from the crystal structure of the selenomethionine derivative of LinD (Semet-LinD) in complex with the natural substrate geraniol, whereas model II (LinD-β-myrcene) was constructed from the crystal structure of LinD in complex with β-myrcene. In addition to a minor conformational difference of the active sites, the two models also differ in the protonation state of key residues. In model I, the pocket residue Asp39' was set to be deprotonated and His129 was protonated on ND1, whereas in model II, Asp38' was set to be deprotonated and His128 was protonated on NE2. Our calculations reveal model II as the active one, which implies that hydration and dehydration are sensitive to the protonation state and fine structure of the active site. On the basis of model II, the conversion details from β-myrcene to geraniol can be obtained. Firstly, β-myrcene is hydrated by a crystal water (W14) and is converted into the stable intermediate (S)-linalool, then linalool is isomerized to geraniol with an overall energy barrier of 24.6 kcal mol-1. Besides, linalool can also reversibly convert into the reactant with an energy barrier of 24.1 kcal mol-1. It is also found that the intermediate IM1 can directly transform to geraniol without first converting to (S)-linalool. His128 and Tyr65 form hydrogen bonds to stabilize the structure of the active site, but they do not act as general acid/base catalysts during the catalytic reactions.
The mechanism of ruthenium-catalyzed
[4 + 1] annulation of benzamide
and propargyl alcohol has been investigated by density functional
theory calculations. The reaction undergoes N–H and C–H
deprotonations by a concerted metalation-deprotonation mechanism to
afford a 5-membered ruthenacyclic species, which then undergoes ring
expansion by alkyne insertion to deliver a 7-membered ring intermediate.
Our study focused on how the successive hydrogen migrations take place
that remains unclear. The 1,2-proton migration and 1,3-proton transfer
from O to C are successively finished by using acetate anion as a
shuttle (a stepwise process). In contrast to the experimental proposal
that the reaction experiences a Ru(II)–Ru(0)–Ru(II)
transformation, our study unveiled a Ru(II)–Ru(IV)–Ru(II)
transformation in the reaction. In addition, our calculations suggested
that the EtO–N bond cleavage rather than the C–H activation
is likely to be the rate-determining step for the entire reaction,
which is not in contradiction with the experimentally reported kinetic
isotope effect values.
In order to explore the inhibitory mechanism of coumarins toward aldose reductase (ALR2), AutoDock and Gromacs software were used for docking and molecular dynamics studies on 14 coumarins (CM) and ALR2 protease. The docking results indicate that residues TYR48, HIS110, and TRP111 construct the active pocket of ALR2 and, besides van der Waals and hydrophobic interaction, CM mainly interact with ALR2 by forming hydrogen bonds to cause inhibitory behavior. Except for CM1, all the other coumarins take the lactone part as acceptor to build up the hydrogen bond network with active-pocket residues. Unlike CM3, which has two comparable binding modes with ALR2, most coumarins only have one dominant orientation in their binding sites. The molecular dynamics calculation, based on the docking results, implies that the orientations of CM in the active pocket show different stabilities. Orientation of CM1 and CM3a take an unstable binding mode with ALR2; their conformations and RMSDs relative to ALR2 change a lot with the dynamic process. While the remaining CM are always hydrogen-bonded with residues TYR48 and HIS110 through the carbonyl O atom of the lactone group during the whole process, they retain the original binding mode and gradually reach dynamic equilibrium.
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