We propose a quantitative structure–property relationship (QSPR) model for prediction of spectral tuning in cyan, green, orange, and red fluorescent proteins, which are engineered by motifs of the green fluorescent protein. Protein variants, in which their chromophores are involved in the π-stacking interaction with amino acid residues tyrosine, phenylalanine, and histidine, are prospective markers useful in bioimaging and super-resolution microscopy. In this work, we constructed training sets of the π-stacked complexes of four fluorescent protein chromophores (of the green, orange, red, and cyan series) with various substituted benzenes and imidazoles and tested the use of dipole moment variation upon excitation (DMV) as a descriptor to evaluate the vertical excitation energies in these systems. To validate this approach, we computed and analyzed electron density distributions of the π-stacked complexes and correlated the QSPR predictions with the reference values of the transition energies obtained using the high-level ab initio quantum chemistry methods. According to our results, the use of the DMV descriptor allows one to predict excitation energies in the π-stacked complexes with errors not exceeding 0.1 eV, which makes this model a practically useful tool in the development of efficient fluorescent markers for in vivo imaging.
We report the results of calculations of the Gibbs energy profiles of the guanosine triphosphate (GTP) hydrolysis by the Arl3-RP2 protein complex using molecular dynamics (MD) simulations with ab initio type QM/MM potentials. The chemical reaction of GTP hydrolysis to guanosine diphosphate (GDP) and inorganic phosphate (Pi) is catalyzed by GTPases, the enzymes, which are responsible for signal transduction in live cells. A small GTPase Arl3, catalyzing the GTP → GDP reaction in complex with the activating protein RP2, constitute an essential part of the human vision cycle. To simulate the reaction mechanism, a model system is constructed by motifs of the crystal structure of the Arl3-RP2 complexed with a substrate analog. After selection of reaction coordinates, energy profiles for elementary steps along the reaction pathway GTP + H2O → GDP + Pi are computed using the umbrella sampling and umbrella integration procedures. QM/MM MD calculations are carried out, interfacing the molecular dynamics program NAMD and the quantum chemistry program TeraChem. Ab initio type QM(DFT)/MM potentials are computed with atom-centered basis sets 6-31G** and two hybrid functionals (PBE0-D3 and ωB97x-D3) of the density functional theory, describing a large QM subsystem. Results of these simulations of the reaction mechanism are compared to those obtained with QM/MM calculations on the potential energy surface using a similar description of the QM part. We find that both approaches, QM/MM and QM/MM MD, support the mechanism of GTP hydrolysis by GTPases, according to which the catalytic glutamine side chain (Gln71, in this system) actively participates in the reaction. Both approaches distinguish two parts of the reaction: the cleavage of the phosphorus-oxygen bond in GTP coupled with the formation of Pi, and the enzyme regeneration. Newly performed QM/MM MD simulations confirmed the profile predicted in the QM/MM minimum energy calculations, called here the pathway-I, and corrected its relief at the first elementary step from the enzyme–substrate complex. The QM/MM MD simulations also revealed another mechanism at the part of enzyme regeneration leading to pathway-II. Pathway-II is more consistent with the experimental kinetic data of the wild-type complex Arl3-RP2, whereas pathway-I explains the role of the mutation Glu138Gly in RP2 slowing down the hydrolysis rate.
The molecular toxicity of uranyl ion (UO22+) in living cells is mainly conditioned by its high affinity to both native and potential metal-binding sites frequently occurring in biomolecules structure. Recent advances in computational and experimental research shed light on the structural properties and functional impacts of uranyl binding to proteins, organic ligands, nucleic acids and their complexes. In the present work, we report the results of the theoretical investigation of the uranyl-mediated loss of DNA-binding activity of PARP-1, eukaryotic enzyme that participates in DNA reparation, cell differentiation, induction of inflammation, etc. Latest experimental studies showed that uranyl ion directly interacts with its DNA-binding subdomains - zinc fingers Zn1 and Zn2, - and changes their tertiary structure. Here, we propose an atomistic mechanism underlying this process and compute the free energy change along the suggested pathway to prove its relevance. According to the results of our QM/MM simulations of Zn2-UO22+complex, uranyl ion replaces zinc in its native binding site, but the corresponding state is destroyed because of the following spontaneous internal hydrolysis of the U-Cys162 coordination bond. Although the enthalpy of hydrolysis is +2.8 kcal/mol, the final value of the free energy of the reaction constitutes -0.6 kcal/mol, due to structure loosening evidenced by solvation and configuration thermodynamic properties calculated using GIST- and MIST-based trajectory processing techniques. The subsequent reorganization of the binding site includes association of uranyl ion with the Glu190/Asp191 acidic cluster and significant perturbations in the domain's tertiary structure, which further decreases the free energy of the non-functional state by 6.8 kcal/mol. The disruption of the DNA-binding interface revealed in our computational simulations is consistent with previous experimental findings and appears to be associated with the loss of the Zn2 affinity for nucleic acids.
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