A multiscale approach to predict the binding mode of metallo beta-lactamase inhibitors.
TL;DR: A reliable computational pipeline is demonstrated that can be applied to inhibitor design for MBLs and other zinc‐metalloenzyme systems and shows the limitations of empirical models for treating these systems and indicates the need for higher level calculations, for example, DFT/MM, for reliable structural predictions.
Abstract: Antibiotic resistance is a major threat to global public health. β-lactamases, which catalyze breakdown of β-lactam antibiotics, are a principal cause. Metallo β-lactamases (MBLs) represent a particular challenge because they hydrolyze almost all β-lactams and to date no MBL inhibitor has been approved for clinical use. Molecular simulations can aid drug discovery, e.g. predicting inhibitor complexes, but empirical molecular mechanics (MM) methods often perform poorly for metalloproteins. Here we present a multiscale approach to model thiol inhibitor binding to IMP-1, a clinically important MBL containing two catalytic zinc ions, and predict the binding mode of a 2-mercaptomethyl thiazolidine (MMTZ) inhibitor. Inhibitors were first docked into the IMP-1 active site, testing different docking programs and scoring functions on multiple crystal structures. Complexes were then subjected to molecular dynamics (MD) simulations and subsequently refined through QM/MM optimization with a density functional theory (DFT) method, B3LYP/6-31G(d), increasing the accuracy of the method with successive steps. This workflow was tested on two IMP-1:MMTZ complexes, for which it reproduced crystallographically observed binding, and applied to predict the binding mode of a third MMTZ inhibitor for which a complex structure was crystallographically intractable. We also tested a 12-6-4 non-bonded interaction model in MD simulations and optimization with a SCC-DFTB QM/MM approach. The results show the limitations of empirical models for treating these systems and indicate the need for higher level calculations, e.g. DFT/MM, for reliable structural predictions. This work demonstrates a reliable computational pipeline that can be applied to inhibitor design for MBLs and other zinc-metalloenzyme systems. This article is protected by copyright. All rights reserved.
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TL;DR: In this paper, the crystal structure of Bacteroides fragilis metallo-β-lactamase was determined by using multiwavelength anomalous diffraction at the zinc absorption edge and refined to 1.85 A resolution.
Abstract: Background: The metallo-β-lactamase from Bacteroides fragilis hydrolyzes a wide range of β-lactam antibiotics, and is not clinically susceptible to any known β-lactamase inhibitors. B. fragilis is associated with post-surgery hospital infections, and there has been a recent report of plasmid-mediated dissemination of the enzyme. Effective inhibitors are therefore urgently needed. Knowledge of the three-dimensional structure will aid in the drug design effort. Results The crystal structure of the enzyme has been determined by using multiwavelength anomalous diffraction at the zinc absorption edge and refined to 1.85 A resolution. The structure is a four-layer α /β/β/ α molecule. The active site, found at the edge of the β sandwich, contains a binuclear zinc center with several novel features. One zinc is tetrahedrally coordinated, the other has a trigonal bipyramidal coordination; a water/hydroxide molecule serves as a ligand for both metals. The residues that coordinate the two zincs are invariant in all metallo-β-lactamases that have been sequenced, except for two conservative replacements. Despite the existence of the pattern for binuclear zinc binding, the reported structure of the Bacillus cereus enzyme contains only a single zinc. Conclusion Structural analysis indicates that affinity for the penta-coordinated zinc can be modulated by neighboring residues, perhaps explaining the absence of the second zinc in the B. cereus structure. Models of bound substrates suggest that the active-site channel can accommodate a wide variety of β-lactams. We propose that the zinc cluster prepares an hydroxide, probably the hydroxide that ligates both zincs, for nucleophilic attack on the carbonyl carbon atom of the β-lactam. The resulting negatively charged tetrahedral intermediate implicated in catalysis is stabilized by an oxyanion hole formed by the side chain of the invariant Asn193 and the tetrahedral zinc.
79 citations
Journal Article•
TL;DR: The parameters presented in this work provide a valuable resource for the molecular simulation community, as they extend the range of metal ions that can be studied using classical approaches, while also providing a starting point for subsequent parametrization of new metal centers.
Abstract: The cationic dummy atom approach provides a powerful nonbonded description for a range of alkaline-earth and transition-metal centers, capturing both structural and electrostatic effects. In this work we refine existing literature parameters for octahedrally coordinated Mn²⁺, Zn²⁺, Mg²⁺, and Ca²⁺, as well as providing new parameters for Ni²⁺, Co²⁺, and Fe²⁺. In all the cases, we are able to reproduce both M²⁺–O distances and experimental solvation free energies, which has not been achieved to date for transition metals using any other model. The parameters have also been tested using two different water models and show consistent performance. Therefore, our parameters are easily transferable to any force field that describes nonbonded interactions using Coulomb and Lennard-Jones potentials. Finally, we demonstrate the stability of our parameters in both the human and Escherichia coli variants of the enzyme glyoxalase I as showcase systems, as both enzymes are active with a range of transition metals. The parameters presented in this work provide a valuable resource for the molecular simulation community, as they extend the range of metal ions that can be studied using classical approaches, while also providing a starting point for subsequent parametrization of new metal centers.
18 citations
TL;DR: In this paper, the authors evaluate the ability of alternative treatments, using (nonbonded) molecular mechanics (MM) and quantum mechanics/molecular mechanics (QM/MM) at semi-empirical (DFTB3) and density functional theory (DFT) levels of theory, to describe the zinc centers of ligand complexes of six metalloenzyme systems differing in coordination geometries, zinc stoichiometries (mono- and dinuclear), and the nature of interacting groups (specifically the presence of zinc-sulfur interactions).
Abstract: Zinc metalloproteins are ubiquitous, with protein zinc centers of structural and functional importance, involved in interactions with ligands and substrates and often of pharmacological interest. Biomolecular simulations are increasingly prominent in investigations of protein structure, dynamics, ligand interactions, and catalysis, but zinc poses a particular challenge, in part because of its versatile, flexible coordination. A computational workflow generating reliable models of ligand complexes of biological zinc centers would find broad application. Here, we evaluate the ability of alternative treatments, using (nonbonded) molecular mechanics (MM) and quantum mechanics/molecular mechanics (QM/MM) at semiempirical (DFTB3) and density functional theory (DFT) levels of theory, to describe the zinc centers of ligand complexes of six metalloenzyme systems differing in coordination geometries, zinc stoichiometries (mono- and dinuclear), and the nature of interacting groups (specifically the presence of zinc-sulfur interactions). MM molecular dynamics (MD) simulations can overfavor octahedral geometries, introducing additional water molecules to the zinc coordination shell, but this can be rectified by subsequent semiempirical (DFTB3) QM/MM MD simulations. B3LYP/MM geometry optimization further improved the accuracy of the description of coordination distances, with the overall effectiveness of the approach depending upon factors, including the presence of zinc-sulfur interactions that are less well described by semiempirical methods. We describe a workflow comprising QM/MM MD using DFTB3 followed by QM/MM geometry optimization using DFT (e.g., B3LYP) that well describes our set of zinc metalloenzyme complexes and is likely to be suitable for creating accurate models of zinc protein complexes when structural information is more limited.
9 citations
TL;DR: In this article , pyrazolone derivatives were used to restore colistin susceptibility of mcr-1-producing bacteria, and the results showed that the pyrazolate with the lowest predicted binding energy (ST3f) was more effective than the other derivatives.
Abstract: The polymyxin colistin is a last line antibiotic for extensively resistant Gram-negative bacteria. Colistin binding to lipid A disrupts the Gram-negative outer membrane, but mobile colistin resistance (mcr) gene family members confer resistance by catalyzing phosphoethanolamine (PEA) transfer onto lipid A, neutralizing its negative charge to reduce colistin interactions. Multiple mcr isoforms have been identified in clinical and environmental isolates, with mcr-1 being the most widespread and mcr-3 being common in South and East Asia. Preliminary screening revealed that treatment with pyrazolones significantly reduced mcr-1, but not mcr-3, mediated colistin resistance. Molecular dynamics (MD) simulations of the catalytic domains of MCR-1 and a homology model of MCR-3, in different protonation states of active site residues H395/H380 and H478/H463, indicate that the MCR-1 active site has greater water accessibility than MCR-3, but that this is less influenced by changes in protonation. MD-optimized structures of MCR-1 and MCR-3 were used in virtual screening of 20 pyrazolone derivatives. Docking of these into the MCR-1/MCR-3 active sites identifies common residues likely to be involved in protein–ligand interactions, specifically the catalytic threonine (MCR-1 T285, MCR-3 T277) site of PEA addition, as well as differential interactions with adjacent amino acids. Minimal inhibitory concentration assays showed that the pyrazolone with the lowest predicted binding energy (ST3f) restores colistin susceptibility of mcr-1, but not mcr-3, expressing Escherichia coli. Thus, simulations indicate differences in the active site structure between MCR-1 and MCR-3 that may give rise to differences in pyrazolone binding and so relate to differential effects upon producer E. coli. This work identifies pyrazolones as able to restore colistin susceptibility of mcr-1-producing bacteria, laying the foundation for further investigations of their activity as phosphoethanolamine transferase inhibitors as well as of their differential activity toward mcr isoforms.
2 citations
References
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TL;DR: A new density functional of the generalized gradient approximation (GGA) type for general chemistry applications termed B97‐D is proposed, based on Becke's power‐series ansatz from 1997, and is explicitly parameterized by including damped atom‐pairwise dispersion corrections of the form C6 · R−6.
Abstract: A new density functional (DF) of the generalized gradient approximation (GGA) type for general chemistry applications termed B97-D is proposed. It is based on Becke's power-series ansatz from 1997 and is explicitly parameterized by including damped atom-pairwise dispersion corrections of the form C(6) x R(-6). A general computational scheme for the parameters used in this correction has been established and parameters for elements up to xenon and a scaling factor for the dispersion part for several common density functionals (BLYP, PBE, TPSS, B3LYP) are reported. The new functional is tested in comparison with other GGAs and the B3LYP hybrid functional on standard thermochemical benchmark sets, for 40 noncovalently bound complexes, including large stacked aromatic molecules and group II element clusters, and for the computation of molecular geometries. Further cross-validation tests were performed for organometallic reactions and other difficult problems for standard functionals. In summary, it is found that B97-D belongs to one of the most accurate general purpose GGAs, reaching, for example for the G97/2 set of heat of formations, a mean absolute deviation of only 3.8 kcal mol(-1). The performance for noncovalently bound systems including many pure van der Waals complexes is exceptionally good, reaching on the average CCSD(T) accuracy. The basic strategy in the development to restrict the density functional description to shorter electron correlation lengths scales and to describe situations with medium to large interatomic distances by damped C(6) x R(-6) terms seems to be very successful, as demonstrated for some notoriously difficult reactions. As an example, for the isomerization of larger branched to linear alkanes, B97-D is the only DF available that yields the right sign for the energy difference. From a practical point of view, the new functional seems to be quite robust and it is thus suggested as an efficient and accurate quantum chemical method for large systems where dispersion forces are of general importance.
23,058 citations
TL;DR: AutoDock Vina achieves an approximately two orders of magnitude speed‐up compared with the molecular docking software previously developed in the lab, while also significantly improving the accuracy of the binding mode predictions, judging by tests on the training set used in AutoDock 4 development.
Abstract: AutoDock Vina, a new program for molecular docking and virtual screening, is presented. AutoDock Vina achieves an approximately two orders of magnitude speed-up compared with the molecular docking software previously developed in our lab (AutoDock 4), while also significantly improving the accuracy of the binding mode predictions, judging by our tests on the training set used in AutoDock 4 development. Further speed-up is achieved from parallelism, by using multithreading on multicore machines. AutoDock Vina automatically calculates the grid maps and clusters the results in a way transparent to the user.
20,059 citations
"A multiscale approach to predict th..." refers methods in this paper
...from PLANTS derives from the combination of ChemScore and Piecewise Linear Potential (PLP) and includes a specific term to evaluate the coordination interaction,(57) while the AutoDock Vina Energy was calculated from the AutoDock4 scoring function and does not include specific terms for metal coordination.(56) Both these tools were tested with different settings in order to determine the setup that most reliably reproduced the crystal structures of protein-ligand...
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TL;DR: A general Amber force field for organic molecules is described, designed to be compatible with existing Amber force fields for proteins and nucleic acids, and has parameters for most organic and pharmaceutical molecules that are composed of H, C, N, O, S, P, and halogens.
Abstract: We describe here a general Amber force field (GAFF) for organic molecules. GAFF is designed to be compatible with existing Amber force fields for proteins and nucleic acids, and has parameters for most organic and pharmaceutical molecules that are composed of H, C, N, O, S, P, and halogens. It uses a simple functional form and a limited number of atom types, but incorporates both empirical and heuristic models to estimate force constants and partial atomic charges. The performance of GAFF in test cases is encouraging. In test I, 74 crystallographic structures were compared to GAFF minimized structures, with a root-mean-square displacement of 0.26 A, which is comparable to that of the Tripos 5.2 force field (0.25 A) and better than those of MMFF 94 and CHARMm (0.47 and 0.44 A, respectively). In test II, gas phase minimizations were performed on 22 nucleic acid base pairs, and the minimized structures and intermolecular energies were compared to MP2/6-31G* results. The RMS of displacements and relative energies were 0.25 A and 1.2 kcal/mol, respectively. These data are comparable to results from Parm99/RESP (0.16 A and 1.18 kcal/mol, respectively), which were parameterized to these base pairs. Test III looked at the relative energies of 71 conformational pairs that were used in development of the Parm99 force field. The RMS error in relative energies (compared to experiment) is about 0.5 kcal/mol. GAFF can be applied to wide range of molecules in an automatic fashion, making it suitable for rational drug design and database searching.
13,615 citations
TL;DR: Together, these backbone and side chain modifications (hereafter called ff14SB) not only better reproduced their benchmarks, but also improved secondary structure content in small peptides and reproduction of NMR χ1 scalar coupling measurements for proteins in solution.
Abstract: Molecular mechanics is powerful for its speed in atomistic simulations, but an accurate force field is required. The Amber ff99SB force field improved protein secondary structure balance and dynamics from earlier force fields like ff99, but weaknesses in side chain rotamer and backbone secondary structure preferences have been identified. Here, we performed a complete refit of all amino acid side chain dihedral parameters, which had been carried over from ff94. The training set of conformations included multidimensional dihedral scans designed to improve transferability of the parameters. Improvement in all amino acids was obtained as compared to ff99SB. Parameters were also generated for alternate protonation states of ionizable side chains. Average errors in relative energies of pairs of conformations were under 1.0 kcal/mol as compared to QM, reduced 35% from ff99SB. We also took the opportunity to make empirical adjustments to the protein backbone dihedral parameters as compared to ff99SB. Multiple sm...
6,367 citations
"A multiscale approach to predict th..." refers methods in this paper
...The ligands were prepared in Antechamber with AM1-BCC charges and GAFF as force field,(44) for which benchmarking in agreement with quantum calculation has been reported,(45) while the protein was parameterized using the ff14SB force field.(46) Given the challenges associated with evaluating the interactions between the zinc ions and...
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TL;DR: The functional classification scheme updated herein is based on the 1995 proposal and includes group 1 (class C) cephalosporinases; group 2 (classes A and D) broad-spectrum, inhibitor-resistant, and extended-spectrums β-lactamases and serine carbapenemases; and group 3 metallo-β-lacticamases.
Abstract: Two classification schemes for β-lactamases are currently in use. The molecular classification is based on the amino acid sequence and divides β-lactamases into class A, C, and D enzymes which utilize serine for β-lactam hydrolysis and class B metalloenzymes which require divalent zinc ions for substrate hydrolysis. The functional classification scheme updated herein is based on the 1995 proposal by Bush et al. (K. Bush, G. A. Jacoby, and A. A. Medeiros, Antimicrob. Agents Chemother. 39:1211-1233, 1995). It takes into account substrate and inhibitor profiles in an attempt to group the enzymes in ways that can be correlated with their phenotype in clinical isolates. Major groupings generally correlate with the more broadly based molecular classification. The updated system includes group 1 (class C) cephalosporinases; group 2 (classes A and D) broad-spectrum, inhibitor-resistant, and extended-spectrum β-lactamases and serine carbapenemases; and group 3 metallo-β-lactamases. Several new subgroups of each of the major groups are described, based on specific attributes of individual enzymes. A list of attributes is also suggested for the description of a new β-lactamase, including the requisite microbiological properties, substrate and inhibitor profiles, and molecular sequence data that provide an adequate characterization for a new β-lactam-hydrolyzing enzyme.
1,878 citations
"A multiscale approach to predict th..." refers background in this paper
...latter mechanism is one of the most important and is most often seen in the form of the hydrolysis of β-lactam antibiotics, catalyzed by serine-β-lactamases (SBLs) and metallo-β-lactamases (MBLs).(3,4) One...
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