1,720,983 research outputs found
First-Principles Modeling of Biological Systems and Structure-Based Drug-Design
No full textMolecular modeling techniques play a relevant role in drug design providing detailed information at atomistic level on the structural, dynamical, mechanistic and electronic properties of biological systems involved in diseases’ onset, integrating and supporting commonly used experimental approaches. These information are often not accessible to the experimental techniques taken singularly, but are of crucial importance for drug design. Due to the enormous increase of the computer power in the last decades, quantum mechanical (QM) or first-principles-based methods have become often used to address biological issues of pharmaceutical relevance, providing relevant information for drug design. Due to their complexity and their size biological systems are often investigated by means of a mixed quantum-classical (QM/MM) approach, which treats at an accurate QM level a limited chemically relevant portion of the system and at the molecular mechanics (MM) level the remaining of the biomolecule and its environment. This method provides a good compromise between computational cost and accuracy, allowing to characterize the properties of the biological system and the (free) energy landscape of the process in study with the accuracy of a QM description. In this review, after a brief introduction of QM and QM/MM methods, we will discuss few representative examples, taken from our work, of the application of these methods in the study of metallo-enzymes of pharmaceutical interest, of metal-containing anticancer drugs targeting the DNA as well as of neurodegenerative diseases. The information obtained from these studies may provide the basis for a rationale structure-based drug design of new and more efficient inhibitors or drugs
The Structural Role of Mg2+ Ions in a Class I RNA Polymerase Ribozyme. A Molecular Simulation Study
No full textAccording to the RNA world hypothesis, self-replicating ribozymes, storing the genetic information and being able to perform catalysis, were the constituents of the first living organisms. In particular, RNA polymerase ribozymes, similar to current proteinaceous enzymatic polymerases, may have been able to promote the synthesis of RNA strands in a primitive world. Polymerase catalysis is usually assisted by Mg2+ ions, but it is not always trivial to find out experimentally the number of Mg2+ ions placed in the active site as well as the identity and the number of their coordination ligands. Here, we addressed this issue in an artificial class I ligase ribozyme. On the basis of a recently solved crystal structure, we constructed computational models of reactant and product states of this ribozyme, considering monometallic and bimetallic species. Our models were relaxed by force field based molecular dynamics (MD) simulations and mixed quantum-classical (QM/MM) MD. The structural and dynamical properties of these models were consistent with experimental data and were validated by a comparison with the catalytic sites of proteinaceous DNA and RNA polymerases. Consistently with enzymatic polymerases, our results suggest that class I RNA ligases most probably contain two magnesium ions in the active site and they may, therefore, catalyze the junction of two RNA strands via "a two Mg2+ ions" mechanism
Influence of the membrane lipophilic environment on the structure and on the substrate access/egress routes of the human aromatase enzyme. A computational study
No full textHuman aromatase (HA), an enzyme located on the membrane of the endoplasmatic reticulum, is of crucial biological importance in the biosynthesis of estrogens. High levels of estrogens are related with important pathologies, conferring to HA a key role as a pharmacological target. In this study we provide, for the first time, an atomistic model of HA embedded on a membrane model to understand the influence of the membrane lipophilic environment on the structural and dynamical properties of HA and on the access/egress pathways of the substrate (androstenedione, ASD) and of the oxygen molecule (involved in the enzymatic process) into/from the HA active site. To this end we used several computational techniques such as force field-based molecular dynamics (MD) simulations, Random Expulsion MD, Steered MD, and Implicit Ligand Sampling. Our results show that the membrane anchoring does not markedly affect the structural properties and the flexibility of the protein, but they clearly point out that the membrane has a marked effect on the access/egress routes of the reactants, stabilizing the formation of different channels for both ASD and O-2 with respect to those observed in pure water solution. Due to the importance of HA in medicine and since access/egress channels may influence its substrate selectivity, a detailed understanding of the role of the membrane in shaping these channels may be of valuable help in drug design
Theoretical Studies of Homogeneous Catalysts Mimicking Nitrogenase
Full text linkThe conversion of molecular nitrogen to ammonia is a key biological and chemical process and represents one of the most challenging topics in chemistry and biology. In Nature the Mo-containing nitrogenase enzymes perform nitrogen ‘fixation’ via an iron molybdenum cofactor (FeMo-co) under ambient conditions. In contrast, industrially, the Haber-Bosch process reduces molecular nitrogen and hydrogen to ammonia with a heterogeneous iron catalyst under drastic conditions of temperature and pressure. This process accounts for the production of millions of tons of nitrogen compounds used for agricultural and industrial purposes, but the high temperature and pressure required result in a large energy loss, leading to several economic and environmental issues. During the last 40 years many attempts have been made to synthesize simple homogeneous catalysts that can activate dinitrogen under the same mild conditions of the nitrogenase enzymes. Several compounds, almost all containing transition metals, have been shown to bind and activate N2 to various degrees. However, to date Mo(N2)(HIPTN)3N with (HIPTN)3N= hexaisopropyl-terphenyl-triamidoamine is the only compound performing this process catalytically. In this review we describe how Density Functional Theory calculations have been of help in elucidating the reaction mechanisms of the inorganic compounds that activate or fix N2. These studies provided important insights that rationalize and complement the experimental findings about the reaction mechanisms of known catalysts, predicting the reactivity of new potential catalysts and helping in tailoring new efficient catalytic compounds
Can multiscale simulations unravel the function of metallo-enzymes to improve knowledge-based drug discovery?
No full textMetallo-enzymes are a large class of biomolecules promoting specialized chemical reactions. Quantum-classical quantum mechanics/molecular mechanics molecular dynamics, describing the metal site at quantum mechanics level, while accounting for the rest of system at molecular mechanics level, has an accessible time-scale limited by its computational cost. Hence, it must be integrated with classical molecular dynamics and enhanced sampling simulations to disentangle the functions of metallo-enzymes. In this review, we provide an overview of these computational methods and their capabilities. In particular, we will focus on some systems such as CYP19A1 a Fe-dependent enzyme involved in estrogen biosynthesis, and on Mg2+-dependent DNA/RNA processing enzymes/ribozymes and the spliceosome, a protein-directed ribozyme. This information may guide the discovery of drug-like molecules and genetic manipulation tools
Sodium-Galactose Transporter: first steps of transport mechanism investigated by molecular dynamics
No full textStructural Role of Uracil DNA Glycosylase for the Recognition of Uracil in DNA Duplexes. Clues from Atomistic Simulations
No full textIn the first stage of the base excision repair pathway the enzyme
uracil DNA glycosylase (UNG) recognizes and excises uracil (U) from DNA
filaments. U repair is believed to occur via a multistep base-flipping process,
through which the damaged U base is initially detected and then engulfed into
the enzyme active site, where it is cleaved. The subtle recognition mechanism
by which UNG discriminates between U and the other similar pyrimidine
nucleobases is still a matter of active debate. Detailed structural information on
the different steps of the base-flipping pathway may provide insights on it.
However, to date only two intermediates have been trapped crystallographically thanks to chemical modifications of the target and/or of its
complementary base. Here, we performed force-field based molecular
dynamics (MD) simulations to explore the structural and dynamical properties
of distinct UNG/dsDNA adducts, containing A:U, A:T, G:U, or G:C base
pairs, at different stages of the base-flipping pathway. Our simulations reveal that if U is present in the DNA sequence a shortlived extra-helical (EH) intermediate exists. This is stabilized by a water-mediated H-bond network, which connects U with
His148, a residue pointed out by mutational studies to play a key role for U recognition and catalysis. Moreover, in this EH
intermediate, UNG induces a remarkable overall axis bend to DNA. We believe this aspect may facilitate the flipping of U, with
respect to other similar nucleobases, in the latter part of the base-extrusion process. In fact, a large DNA bend has been
demonstrated to be associated with a lowering of the free energy barrier for base-flipping. A detailed comparison of our results
with partially flipped intermediates identified crystallographically or computationally for other base-flipping enzymes allows us to
validate our results and to formulate hypothesis on the recognition mechanism of UNG. Our study provides a first ground for a
detailed understanding of the UNG repair pathway, which is necessary to devise new pharmaceutical strategies for targeting
DNA-related pathologies
Single or multiple access channels to the CYP450s active site? An answer from free energy simulations of the human aromatase enzyme
No full textCytochromes P450 (CYP450s), in particular, CYP19A1 and CYP17A1, are key clinical targets of breast and prostate anticancer therapies, critical players in drug metabolism, and their overexpression in tumors is associated with drug resistance. In these enzymes, ligand (substrates, drugs) metabolism occurs in deeply buried active sites accessible only via several grueling channels, whose exact biological role remains unclear. Gaining direct insights on the mechanism by which ligands travel in and out is becoming increasingly important given that channels are involved in the modulation of binding/dissociation kinetics and the specificity of ligands toward a CYP450. This has profound implications for enzymatic efficiency and drug efficacy/toxicity. Here, by applying free energy methods, for a cumulative simulation time of 20 s, we provide detailed atomistic characterization and free energy profiles of the entry/exit routes preferentially followed by a substrate (androstenedione) and a last-generation inhibitor (letrozole) to/from the catalytic site of CYP19A1 (the human aromatase (HA) enzyme), a key clinical target against breast cancer, studied here as prototypical CYP450. Despite the remarkably different size/shape/hydrophobicity of the ligands, two channels appear accessible to their entrance, while only one exit route appears to be preferential. Our study shows that the preferential paths may be conserved among different CYP450s. Moreover, our results highlight that, at least in the case of HA, ligand channeling is associated with large enzyme structural rearrangements. A wise choice of the computational method and very long simulations are, thus, required to obtain fully converged quantitative free energy profiles, which might be used to design novel biocatalysts or next-generation cytochrome inhibitors with an in silico tuned K-m
Computational Approaches Elucidate the Allosteric Mechanism of Human Aromatase Inhibition: A Novel Possible Route to Small-Molecule Regulation of CYP450s Activities?
No full textHuman aromatase (HA) is a P450 cytochrome (CYP) with an essential role in estrogen biosynthesis. Since more than 70% of breast cancers are positive for estrogenic receptor (ER), the reduction of estrogen physiological concentrations through HA inhibition is one of most important therapeutic strategies against this cancer type. Recently, experimental evidence showed that selected taxmoxifen metabolites, which are typically used as estrogen receptor modulators (SERMs), inhibit HA through an allosteric mechanism. In this work, we present a computational protocol to (i) characterize the structural framework and (ii) define the atomistic details of the determinants for the noncompetitive inhibition mechanism. Our calculations identify two putative binding sites able to efficiently bind all tamoxifen metabolites. Analysis of long-scale molecular dynamics simulations reveal that endoxifen, the most effective noncompetitive inhibitor, induces significant enzyme rigidity by binding in one of the possible peripheral sites. The consequence of this binding event is the suppression of one of the functional enzymatic collective motions associated with breathing of the substrate access channel. Moreover, an internal dynamics-based alignment of HA with six other human cytochromes shows that this collective motion is common to other members of the CYP450 protein family. On this basis, our findings may thus be of help for the development of new (pan)inhibitors for the therapeutic treatment of cancer, targeting and modulating the activity of HA and of estrogen receptor, and may also stimulate the development of new drug design strategies for chemoprevention and chemoprotection via allosteric inhibition of CYP450 protein
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