On the Catalytic Activity of the Engineered Coiled-Coil Heptamer Mimicking the Hydrolase Enzymes: Insights from a Computational Study

Abstract

Recently major advances were gained on the designed proteins aimed to generate biomolecular mimics of proteases. Although such enzyme-like catalysts must still suffer refinements for improving the catalytic activity, at the moment, they represent a good example of artificial enzymes to be tested in different fields. Herein, a de novo designed homo-heptameric peptide assembly (CC-Hept) where the esterase activity towards p-nitro-phenylacetate was obtained for introduction of the catalytic triad (Cys-His-Glu) into the hydrophobic matrix, is the object of the present combined molecular dynamics and quantum mechanics/molecular mechanics investigation. Constant pH Molecular Dynamics simulations on the apoform of CC-Hept suggested that the Cys residues are present in the protonated form. Molecular dynamics (MD) simulations of the enzyme–substrate complex evidenced the attitude of the enzyme-like system to retain water molecules, necessary in the hydrolytic reaction, in correspondence of the active site, represented by the Cys-His-Glu triad on each of the seven chains, without significant structural perturbations. A detailed reaction mechanism of esterase activity of CC-Hept-Cys-His-Glu was investigated on the basis of the quantum mechanics/molecular mechanics calculations employing a large quantum mechanical (QM) region of the active site. The proposed mechanism is consistent with available esterases kinetics and structural data. The roles of the active site residues were also evaluated. The deacylation phase emerged as the rate-determining step, in agreement with esterase activity of other natural proteases.

The Se–S Bond Formation in the Covalent Inhibition Mechanism of SARS-CoV-2 Main Protease by Ebselen-like Inhibitors: A Computational Study

“…At the end of the activation step, the EI' structure, is characterized by a CysS − /HisH + zwitterionic pair. Irrespective of an HIE or HID form, our calculations for ES therefore support the finding that Cys145 of the M pro of SARS-CoV-2 works in the neutral form and represents the resting state of the enzyme as previously noted in similar systems [71,80].…”

On the Recognition of Natural Substrate CTP and Endogenous Inhibitor ddhCTP of SARS-CoV-2 RNA-Dependent RNA Polymerase: A Molecular Dynamics Study

“…The four most populated structures were used as a starting point for the docking procedure. The described procedure has been successfully adopted in a number of investigations on different enzymes. − …”

“…It was suggested that during the reaction, a shrinking of this distance may occur, leading O3′ to become closer to Pα and consequently facilitating the nucleophilic attack. The little longer distance in ddhCTP than that in CTP could affect the O3′–P α alignment for catalysis during the nucleophilic attack, in the reactive Michaelis–Menten complex. − …”

Structural Characterization of Peripolin and Study of Antioxidant Activity of HMG Flavonoids from Bergamot Fruit

“…The General Amber Force Field (GAFF) and the Restrained Electrostatic Potential method were adopted to extrapolate non-bonding parameters and charges, respectively [ 26 , 27 ]. The here-adopted parametrization procedure represents a well-assessed protocol that was successfully adopted in previous studies [ 28 , 29 , 30 ].…”