Identification of a Cryptic Pocket in Methionine Aminopeptidase-II Using Adaptive Bandit Molecular Dynamics Simulations and Markov State Models.

Methionine aminopeptidase-II (MetAP-II) is a metalloprotease, primarily responsible for the cotranslational removal of the N-terminal initiator methionine from the nascent polypeptide chain during protein synthesis. MetAP-II has been implicated in angiogenesis and endothelial cell proliferation and is therefore considered a validated target for cancer therapeutics. However, there is no effective drug available against MetAP-II. In this study, we employ Adaptive Bandit molecular dynamics simulations to investigate the structural dynamics of the apo and ligand-bound MetAP-II. Our results focus on the dynamic behavior of the disordered loop that is not resolved in most of the crystal structures. Further analysis of the conformational flexibility of the disordered loop reveals a hidden cryptic pocket that is predicted to be potentially druggable. The network analysis indicates that the disordered loop region has a direct signaling route to the active site. These findings highlight a new way to target MetAP-II by designing inhibitors for the allosteric site within this disordered loop region.

Attempting Well-Tempered Funnel Metadynamics Simulations for the Evaluation of the Binding Kinetics of Methionine Aminopeptidase-II Inhibitors.

Understanding the unbinding kinetics of protein-ligand complexes is considered a significant approach for the design of ligands with desired specificity and safety. In recent years, enhanced sampling methods have emerged as effective tools for studying the unbinding kinetics of protein-ligand complexes at the atomistic level. MetAP-II is a target for the treatment of cancer for which not a single effective drug is available yet. The identification of the dissociation rate of ligands from the complexes often serves as a better predictor for in vivo efficacy than the ligands' binding affinity. Here, funnel-based restraint well-tempered metadynamics simulations were applied to predict the residence time of two ligands bound to MetAP-II, along with the ligand association and dissociation mechanism involving the identification of the binding hotspot during ligand egress. The ligand-egressing route revealed by metadynamics simulations also correlated with the identified pathways from the CAVER analysis and by the enhanced sampling simulation using PLUMED. Ligand 1 formed a strong H-bond interaction with GLU364 estimating a higher residence time of 28.22 ± 5.29 ns in contrast to ligand 2 with a residence time of 19.05 ± 3.58 ns, which easily dissociated from the binding pocket of MetAP-II. The results obtained from the simulations were consistent to reveal ligand 1 being superior to ligand 2; however, the experimental data related to residence time were close for both ligands, and no kinetic data were available for ligand 2. The current study could be considered the first attempt to apply an enhanced sampling method for the evaluation of the binding kinetics and thermodynamics of two different classes of ligands to a binuclear metalloprotein.

Deciphering Structural and Dynamical Properties of Hydrated Cobalt Porphyrins via Ab Initio Quantum Mechanical Charge Field Molecular Dynamics Simulation.

The present study successfully implemented the ab initio quantum mechanical charge field molecular dynamics (QMCF MD) formalism for the investigation of structural and dynamical properties of hydrated cobalt-porphyrin complexes. Considering the significance of cobalt ions in biological systems (for instance, vitamin B12), which reportedly incorporate cobalt ions in a d6, low spin, +3 state chelated in the corrin ring, an analog of porphyrin, the current study is focused on cobalt in the oxidation states +2 and +3 bound to the parent porphyrin lead structures embedded in an aqueous solution. These cobalt-porphyrin complexes were investigated in terms of their structural and dynamical properties at the quantum chemical level. The structural attributes of these hydrated complexes revealed the contrasting features of the water binding to these solutes, including a detailed evaluation of the associated dynamics. The study also yielded notable findings in regard to the respective electronic configurations vs coordination, which suggested that Co(II)-POR possesses a 5-fold square pyramidal coordination geometry in an aqueous solution containing the metal ion coordinating to four nitrogen atoms of the porphyrin ring and one axial water as the fifth ligand. On the other hand, high-spin Co(III)-POR was hypothesized to be more stable due to the smaller size-to-charge ratio of the cobalt ion, but the high-spin complex demonstrated unstable structural and dynamical behavior. However, the corresponding properties of the hydrated Co(III)LS-POR revealed a stable structure in an aqueous solution, thus suggesting the Co(III) ion to be in a low-spin state when bound to the porphyrin ring. Moreover, the structural and dynamical data were augmented by computing the free energy of water binding to the cobalt ions and the solvent-accessible surface area, which provide further information on thermochemical properties of the metal-water interaction and the hydrogen bonding potential of the porphyrin ring in these hydrated systems.

Interaction of gentamicin and gentamicin-AOT with poly-(lactide-co-glycolate) in a drug delivery system - density functional theory calculations and molecular dynamics simulation.

Gentamicin is used to treat brucellosis, an infectious disease caused by the Brucella species but the drug faces several issues such as low efficacy, instability, low solubility, and toxicity. It also has a very short half-life, therefore, requiring frequent dosing. Consequently, several other antibiotics are also being used for the treatment of brucellosis as a single dose as well as in combination with other antibiotics but none of these therapies are satisfactory. Nanoparticles in particular polymer-based ones utilizing polymers that are biodegradable and biocompatible for instance PLGA are a method of choice to overcome such drug delivery issues and enable potential targeted delivery. The current study focuses on the evaluation of the structural and dynamical properties of a drug-polymer system consisting of gentamicin drug and PLGA polymer nanoparticles in the water representing a targeted drug delivery system for the treatment of brucellosis. For this purpose, all-atom molecular dynamics simulations were carried out on the drug-polymer systems in the absence and presence of the surfactant bis(2-Ethylhexyl) sulfosuccinate (AOT) to determine the structural and dynamical properties as well as the effect of the surfactant on these properties. We also investigated systems in which the polymer constituents were in the form of monomeric units toward decoupling the primary interactions of the monomer units and polymer effects. The simulation results explain the nature of the interactions between the drug and the polymer as well as transport properties in terms of drug diffusion coefficients, which characterize the molecular behavior of gentamicin-polymer nanoparticles for use in brucellosis.

Structure and dynamics of 5-lipoxygenase and its complexes - a molecular dynamics simulation study.

Molecular dynamics simulations were applied to human 5-LOX to obtain detailed information on its structure and dynamics with and without ligands. The dynamical properties evaluated based on root mean square deviations, root mean square fluctuations and secondary structure prediction helped decipher the contrast dynamic behavior of the systems pointing toward the ligand binding effect. The ligand binding to the protein also perturbed other properties of the protein such as the central bending of the protein and water coordination to the metal ion. The central bending in the protein was reported to be very significant that was associated with the allosteric modulation in the lipoxygenases; therefore, on a similar line, the central bending was evaluated in terms of hinge angle analysis which showed substantial bending between the C-terminal and the N-terminal domain via the linker residues which connects the two domains. On the other hand, the suspected water coordination to the metal ion in the protein was ruled out by computing the iron-water radial distribution function which showed that the water molecule was not found to be in the vicinity of the metal ion. Finally, the binding free energy was estimated for Zileuton and CAPE1 inhibitors bound to 5-LOX via the thermodynamic integration approach which showed that CAPE1 had a strong binding potential for the active site of the protein compared to Zileuton, and the free energy data correlated well with their IC50 values corresponding to the high inhibition potential of CAPE1 compared to Zileuton.

Genomic analysis of Chryseobacterium indologenes and conformational dynamics of the selected DD-peptidase.

Chrysobacterium indologenes is an emerging MDR pathogen that belongs to the family Flavobacteriaceae. The genome of the C. indologenes, isolated from the nephrotic patient, was sequenced through Illumina MiSeq. The pangenomics of available 56 C. indologenes strains using BPGA revealed an open pangenome (n=5553 CDS), core genome (2141), and accessory genome (2013). The CEG/DEG database identified 662 essential genes that drastically reduced to 68 genes after non-homology analyses towards human and gut microbiome. Further filtering the data for other drug target prioritizing parameters resulted in 32 putative targets. Keeping in view the crucial role played in cell wall biosynthesis, dacB was selected as the final target that encodes D-alanyl-d-alanine carboxypeptidase/endopeptidase (DD-peptidase). The 3D structure of dacB was modelled and rendered to docking analyses against two compound libraries of African plants (n=6842) and Tibetan medicines (n=52). The ADMET profiling exhibited the physicochemical properties of final compounds. The MD simulations showed the stability of inhibitor-DD-peptidase complex and interactions in terms of RMSD, RMSF, binding free energy calculation and H-bonding. We propose that the novel compounds Leptopene and ZINC95486338 from our findings might be potent DD-peptidase inhibitors that could aid in the development of new antibiotic-resistant therapy for the emerging MDR C. indologenes.

Interaction of Phthalates with Lipid Bilayer Membranes.

Phthalates are esters of phthalic acid, widely used as additives in the manufacture of plastics. They are not covalently linked to polymer chains and can easily leach out, disperse in the environment, and get into contact with living organisms. Several short chain phthalates are classified as endocrine disruptors or hormonal active agents, and have also been reported to promote various kinds of cancer. However, the biological effects of longer chain analogues are less well known. Moreover, little is known on the permeation of phthalates and their metabolites through biological membranes and on their effects on the physical properties of membranes. Here we explore the interaction of a group of phthalates and their main metabolites with model biological membranes. We focus on three industrially relevant phthalates, with acyl chains of different sizes, and their monoester metabolites. We use molecular dynamics simulations to predict the distribution in model membranes, as well as permeabilities and effects on the structural, dynamic, and elastic properties of the membranes. We find that alterations of membrane properties are significant and only weakly affected by the size of acyl chains, suggesting that modifications of molecular size may not be sufficient to reduce the impact of this class of molecules on the environment and health.

Dynamics of metal binding and mutation in yybP-ykoY riboswitch of Lactococcus lactis.

Riboswitch is a regulatory segment of messenger RNA (mRNA), which by binding to various cellular metabolites regulates the activity of mRNA via modulating transcription, translation, alternative splicing, and stability of the mRNA. yybP-ykoY riboswitch of Lactococcus lactis, which is present upstream of the yoaB gene, functions as a Mn2+-specific genetic ON-switch, and modulates expression of proteins which are significant for Mn2+ homeostasis. The P1.1 switch helix of the aptamer domain of the riboswitch contains an intrinsic transcription terminator structure, which gets stabilized with Mn2+ binding and causes disruption of terminator structure and allows the continuation of transcription. The current research work involved the evaluation of structural and dynamical properties of the yybP-ykoY riboswitch of L. lactis in its Mn2+-free, Mn2+-bound (wild-type), and Mn2+-bound mutant (A41U) states by applying molecular dynamics simulations. Based on the simulations, the effects of Mn2+ absence and A41U mutation were evaluated on the structure and dynamics of the riboswitches followed by the computation of the free energy of metal binding in the wild-type and the mutant riboswitches. The simulation results provided insights into the properties of the riboswitch with the focus on the dynamics of the P1.1 switch helix, and the manganese binding site designated as MB site, as well as the relative stability of the wild-type and the mutant riboswitches, which helped to understand the structural and dynamical role of the metal ion involved in the function of Mn2+-sensing riboswitch.

Investigation of the structural and dynamical properties of human uncoupling protein 2 through molecular dynamics simulations.

Uncoupling protein 2 (UCP2) is an integral membrane protein that belongs to the family of mitochondrial anion carrier proteins. The absence of human UCP2 structure, lack of understanding of Cl- ion transport mechanism in the UCP2 and the associated biological functions motivated us to model the protein and investigate its structural and dynamical properties in a realistic mitochondrial lipid membrane system. The lipid-protein and protein-protein interactions were probed since they were found to be responsible for the conformational changes of the transmembrane (TM) helices which are involved in facilitating Cl- ion transport. Here, we employed multiscale molecular dynamics simulations including unbiased and biased MD for the investigation of the transport pathway in hUCP2 and interactions of the ion with TM helices within a membrane environment. We initially validated the hUCP2 model in the lipid membrane and then explored the transport pathway of Cl- ion and its interaction with positive residues of TM2 helix that have been reported to play a major role in the Cl- ion transport along with other TM helices of the protein. The simulation results suggest that the TM2 helix plays an important role in the formation of a stable ion channel due to the presence of arginine residues, in particular Arg88 which was found to be a key residue to maintain the channel pore through which the movement of Cl- ions occurs. Based on the results, it can be said that the study provides an atomic-level description of the Cl- ion transport mechanism in hUCP2 embedded in the mitochondrial lipid membrane.

Hydration modulates oxygen channel residues for oxygenation of cysteine dioxygenase: Perspectives from molecular dynamics simulations.

Cysteine dioxygenase (CDO) regulates the concentration of l-cysteine substrate by its oxidation in the body to prevent different diseases, including neurodegenerative and autoimmune diseases. CDO catalyzes the oxidation of thiol group of l-cysteine to l-cysteine sulfinic acid using molecular oxygen. In this study, molecular dynamics simulations were applied to ligand-free CDO, cysteine-bound CDO, and oxygen-bound CDO-cysteine complex which were primarily subjected to the evaluation of their structural and dynamical properties. The simulation data provided significant information not only on the conformational changes of the enzyme after its ligation but also on the co-ligation by sequential binding of l-cysteine and molecular oxygen. It was found that the ligation and co-ligation perturbed the active site region as well as the overall protein dynamics which were analyzed in terms of root mean square deviation, root mean square fluctuation and dynamic cross correlation matrices as well as principal component analysis. Furthermore, oxygen transport pathways were successfully explored by taking various tunnel clusters into account and one of those clusters was given preference based on the throughput value. The bottleneck formed by different amino acid residues was examined to figure out their role in the oxygenation process of the enzyme. The residues forming the tunnel's bottleneck and their dynamics mediated by water molecules were further investigated using radial distribution functions which gave insights into the hydration behavior of these residues. The findings based on the hydration behavior in turn served to explore the water-mediated dynamics of these residues in the modulation of the pathway, including tunnel gating for the oxygen entry and diffusion to the active site, which is essential for the CDO's catalytic function.

Inter-Subunit Dynamics Controls Tunnel Formation During the Oxygenation Process in Hemocyanin Hexamers.

Hemocyanin from horseshoe crab in its active form is a homo-hexameric protein. It exists in open and closed conformations when transitioning between deoxygenated and oxygenated states. Here, we present a detailed dynamic atomistic investigation of the oxygenated and deoxygenated states of the hexameric hemocyanin using explicit solvent molecular dynamics simulations. We focus on the variation in solvent cavities and the formation of tunnels in the two conformational states. By employing principal component analysis and CVAE-based deep learning, we are able to differentiate between the dynamics of the deoxy- and oxygenated states of hemocyanin. Finally, our results identify the deoxygenated open conformation, which adopts a stable, closed conformation after the oxygenation process.

Iron coordination to pyochelin siderophore influences dynamics of FptA receptor from Pseudomonas aeruginosa: a molecular dynamics simulation study.

FptA is a TonB-dependent transporter that permits the high affinity binding and transport of Fe(III)-pyochelin complex across the outer membrane of Pseudomonas aeruginosa. Molecular dynamics simulations were employed to FptA receptor and its complexes with pyochelin, and co-crystallized Fe(III)-pyochelin-ethanediol and Fe(III)-pyochelin-water embedded in dilauroyl phosphatidyl choline bilayer for the evaluation of their structural and dynamical properties. The evaluation of properties of the receptor bound to pyochelin molecule and Fe(III)-pyochelin complexes helped to figure out the iron coordination effect on the receptor properties. Moreover, comparison of these four simulation systems revealed further information on the dynamical changes occurred in extracellular loops, in particular loop-7 corresponding to the missing amino acid residues including the close-by loop-8 that was largely affected by the metal coordination to pyochelin. The binding of iron to pyochelin molecule affected the overall structure of the receptor therefore, evaluation fo the gyration radii and hydrogen bonding were evaluated as well as analysis of the pore size were also carried out to understand the effect of metal coordination on the dynamics of the helices which form a kind of translocation channel to transport the siderophore across the FptA protein into the periplasmic space. The properties of each component of the molecular systems were therefore observed to be perturbed by the incorporation of iron to the pyochelin molecule thus demonstrating that the bacteria use its receptor to abstract and transport iron from extracellular environment for its survival and that was made possible to understand at the molecular level through successful implementation of molecular dynamics simulations.

Evaluation of a sesquiterpene as possible drug lead against gelatinases via molecular dynamics simulations.

Malignant tumors can be targeted by accounting for their metastatic capabilities. Matrix metalloproteinases (MMPs) are the key players in tumor metastasis facilitating through their proteolytic activities of angiogenesis and extracellular matrix components (ECM) degradation. MMP-2 and MMP-9 being the members of a distinguished class of MMPs more commonly known as gelatinases are the prominent enzymes which are involved in different cancer progression stages. Targeting these isoforms specifically has always been a challenging task due to highly similar structural and functional features among the other members of MMPs with well preserve active sites containing catalytic zinc atom that was the only reason that none of the MMP inhibitor has been successfully marketed for the tumor pathology up till now. Therefore, non-competitive inhibitors with different structural attributed are needed to be evaluated at the molecular level for further experiments. The present study deals with the application of molecular dynamics simulation for the investigation of an alternative pathway for the inhibition of MMP-2 and MMP-9 by a sesquiterpene isolated from Polygonum barbatum which demonstrates the characteristics binding to the S1' subsite of the enzymes followed by in vitro gene expression studies. The simulation results provide information on the possible binding profile producing inhibitory effects imposed by the inhibitor to these enzymes by acquiring different structural and dynamical features. Moreover, thermodynamic quantities based on the computationally intensive thermodynamic integration approach were also obtained in terms of inhibitor binding affinity computed for the inhibitor against MMP-2 and MMP-9 that completely augmented the experimental gene expression study.Communicated by Ramaswamy H. Sarma.

Exploring interfacial dynamics in homodimeric S-ribosylhomocysteine lyase (LuxS) from Vibrio cholerae through molecular dynamics simulations.

To the best of our knowledge, this is the first molecular dynamics simulation study on the dimeric form of the LuxS enzyme from Vibrio cholerae to evaluate its structural and dynamical properties including the dynamics of the interface formed by the two monomeric chains of the enzyme. The dynamics of the interfacial region were investigated in terms of inter-residual contacts and the associated interface area of the enzyme in its ligand-free and ligand-bound states which produced characteristics contrast in the interfacial dynamics. Moreover, the binding patterns of the two inhibitors (RHC and KRI) to the enzyme forming two different enzyme-ligand complexes were analyzed which pointed towards a varying inhibition potential of the inhibitors as also revealed by the free energies of ligand binding. It is shown that KRI is a more potent inhibitor than RHC - a substrate analogue, showing correlation with experimental data. Moreover, the role of a loop in chain B of the enzyme was found to facilitate the binding of RHC similar to that of the substrate, while KRI demonstrates a differing binding pattern. The computation of the free energy of binding for the two ligands was also carried out via thermodynamic integration which ultimately served to correlate the dynamical properties with the inhibition potential of two different ligands against the enzyme. Furthermore, this successful study provides a rational to suggest novel LuxS inhibitors which could become promising candidates to treat the diseases caused by a broad variety of bacterial species.

Solvation of cholesterol in different solvents: a molecular dynamics simulation study.

To the best of our knowledge, molecular dynamics simulations of an isolated cholesterol immersed in four different solvents of varying polarity, such as water, methanol, dimethyl sulfoxide and benzene, were reported for the first time to gain insights into the structural and dynamical properties. The study was mainly focused on the evaluation of solvation of cholesterol with respect to its hydrophilic and hydrophobic structural components in the form of respective functional groups interacting with the solvents. Structural evaluations suggested that both hydrophilic and hydrophobic groups of cholesterol were interacting with the solvents, in particular methanol and dimethyl sulfoxide, which presented both types of interactions that are polar and non-polar. On the other hand, the highly polar water and non-polar benzene demonstrated extreme solvation behavior, since water was involved only in hydrogen bonding to the solute hydroxyl group and non-polar benzene formed strong van der Waals interactions only. Furthermore, the hydrophobic effect of cholesterol was also analyzed mainly in polar solvents, as the effect was more pronounced in the polar environment thereby preventing the solvent mobility in the solvation layer(s). The dynamical properties in terms of lateral diffusion and hydrogen bond dynamics as well as free energies of solvation also corroborated the findings based on the structural data and the hydrophobic character of cholesterol was later quantified by the computation of the averaged solvent accessible surface area. The polarity effect of the solvents on the aggregation property of cholesterol was further investigated, which is of big concern from the clinical point of view due to its major role in cardiovascular ailments. It was another major finding of the present study that aggregation was shown to be facilitated by highly polar solvents like water.

Hydration of Closely Related Manganese and Magnesium Porphyrins in Aqueous Solutions: Ab Initio Quantum Mechanical Charge Field Molecular Dynamics Simulation Study.

To the best of our knowledge, the current study based on ab initio quantum mechanical charge field molecular dynamics (QMCF-MD) is the first to explore the difference in the hydration behavior between Mn(II)- and Mg(II)-associated porphyrins (Mn(II)-POR and Mg(II)-POR) in aqueous solution. The simulation study highlights similar and dissimilar characteristics of the structural, dynamical, and thermodynamical properties of these closely related metals bound to porphyrins in aqueous solution. The structural analysis is based on radial and angular distribution functions, coordination number distributions, and angular-radial distributions. Both hydrated systems demonstrate similar pentacoordinated structures formed via the axial coordination of one water molecule to the metal ion in addition to the four nitrogen atoms of the porphyrin ring. However, in the case of Mn(II)-POR, the formation of a distorted square pyramidal geometry was observed. It was envisaged as a weak coordination of the water molecule to the Mn(II) atom and thus higher atomic fluctuation for all atoms in contrast to that for the hydrated Mg(II)-POR. The dynamical data in terms of the mean residence times, velocity autocorrelation function, free energy, and other parameters revealed the difference in the metal binding effect because the Mn(II) atom was observed to inhibit H-bond formation more than the presence of Mg(II) atoms in the core of the porphyrin. The current study thus highlights the significant differences in the structural and dynamical properties of Mn(II)- and Mg(II)-associated porphyrin systems.

Highly selective enrichment of phosphopeptides using poly(dibenzo-18-crown-6) as a solid-phase extraction material.

A poly(dibenzo-18-crown-6) was used as a new solid-phase extraction material for the selective enrichment of phosphopeptides. Isolation of phosphopeptides was achieved based on specific ionic interactions between poly(dibenzo-18-crown-6) and the phosphate group of phosphopeptides. Thus, a method was developed and optimized, including loading, washing and elution steps, for the selective enrichment of phosphopeptides. To assess this potential, tryptic digest of three proteins (α- casein, ß-casein and ovalbumin) was applied on poly(dibenzo-18-crown-6). The nonspecific products were removed by centrifugation and washing. The spectrometric analysis was performed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Highly selective enrichment of both mono- and multiphosphorylated peptides was achieved using poly(dibenzo-18-crown-6) as solid-phase extraction material with minimum interference from nonspecific compounds. Furthermore, evaluation of the efficiency of the poly(dibenzo-18-crown-6) was performed by applying the digest of egg white. Finally, quantum mechanical calculations were performed to calculate the binding energies to predict the affinity between poly(dibenzo-18-crown-6) and various ligands. The newly identified solid-phase extraction material was found to be a highly efficient tool for phosphopeptide recovery from tryptic digest of proteins.

Hydration facilitates oxygenation of hemocyanin: perspectives from molecular dynamics simulations.

Molecular dynamics simulations were applied to deoxy- and oxy-hemocyanins using newly developed force field parameters for the dicopper site to evaluate their structural and dynamical properties. Data obtained from the simulations provided information of the oxygenation effect on the active site and overall topology of the protein that was analyzed by root-mean-square deviations, b-factors, and dicopper coordination geometries. Domain I of the protein was found to demonstrate higher flexibility with respect to domain II because of the interfacial rotation between domain I and II that was further endorsed by computing correlative domain movements for both forms of the protein. The oxygenation effect on the overall structure of the protein or polypeptide subunit was further explored via gyration radii evaluated for the metal-binding domain and for the whole subunit. The evaluation of hydration dynamics was carried out to understand the water mediated role of amino acid residues of the solvent tunnel facilitating the entry of oxygen molecule to the dicopper site of hemocyanin.

Hydration of iron-porphyrins: ab initio quantum mechanical charge field molecular dynamics simulation study.

The ab initio quantum mechanical charge field molecular dynamics (QMCF-MD) simulation approach was successfully applied to Fe2+-P and Fe3+-P in water to evaluate their structural, dynamical and energetic properties. Based on the structural data, it was found that Fe2+-P accommodates one water molecule in the first coordination sphere of the Fe2+ ion including the four nitrogen atoms of the porphyrin system coordinating with central metal species. On the other hand, two water molecules were coordinated to Fe3+-P, thus forming a hexa-coordinated species. Comparison of dynamical properties such as the vibrational power spectrum and ligand mean residence times to other metal-free porphyrin systems demonstrate the ions' influence on the hydration structure, enabling a characterisation of the strong interaction of the ions which greatly reduces the hydrogen bonding potential of the complex. The association of water molecules with the metal ions in both solutes was quantified by computing the free energy of binding obtained via the potential of mean force. This further confirmed the strong association of water to the metal ions which was conversely weak as inferred from the energetic data for the Fe2+-P system.

Syntheses of 4,6-dihydroxypyrimidine diones, their urease inhibition, in vitro, in silico, and kinetic studies.

A library of 4,6-dihydroxypyrimidine diones (1-35) were synthesized and evaluated for their urease inhibitory activity. Structure-activity relationships, and mechanism of inhibition were also studied. All compounds were found to be active with IC50 values between 22.6±1.14-117.4±0.73µM, in comparison to standard, thiourea (IC50=21.2±1.3µM). Kinetics studies on the most active compounds 2-7, 16, 17, 28, and 33 were performed to investigate their modes of inhibition, and dissociation constants Ki. Compounds 2, 3, 7, 16, 28, and 33 were found to be mixed-type of inhibitors with Ki values in the range of 7.91±0.024-13.03±0.013µM, whereas, compounds 4-6, and 17 were found to be non-competitive inhibitors with Ki values in the range of 9.28±0.019-13.05±0.023µM. In silico study was also performed, and a good correlation was observed between experimental and docking studies. This study is continuation of our previously reported urease inhibitory activity of pyrimidine diones, representing potential leads for further research as possible treatment of diseases caused by ureolytic bacteria.