Showing papers by "Jeffry D. Madura published in 2010"

Molecular dynamics of leucine and dopamine transporter proteins in a model cell membrane lipid bilayer.

Abstract: The dopamine transporter (DAT) operates via facilitated diffusion, harnessing an inward Na(+) gradient to drive dopamine from the extracellular synaptic cleft to the neuron interior. The DAT is relevant to central nervous system disorders such as Parkinson disease and attention-deficit hyperactivity disorder and is the primary site of action for the abused psychostimulants cocaine and amphetamines. Crystallization of a DAT homolog, the bacterial leucine transporter LeuT, provided the first reliable 3-D DAT template. Here, the LeuT crystal structure and the DAT molecular model have been combined with their respective substrates, leucine and dopamine, in lipid bilayer molecular dynamics simulations toward tracking substrate movement along the protein's substrate/ion permeation pathway. Specifically, movement of residue pairs that comprise the "external gate" was followed as a function of substrate presence. The transmembrane (TM) 1 arginine-TM 10 aspartate strut formed less readily in DAT compared with LeuT, with or without substrate present. For LeuT but not DAT, the addition of substrate enhanced the chances of forming the TM 1-10 bridge. Also, movement of the fourth extracellular loop EL-4 in the presence of substrate was more pronounced for DAT, the EL-4 unwinding to a degree. The overall similarity between the LeuT and DAT molecular dynamics simulations indicated that LeuT was a legitimate model to guide DAT structure-function predictions. There were, nevertheless, differences significant enough to allow for DAT-unique insights, which may include how cocaine, methylphenidate (Ritalin, NIDA Drug Supply, Rockville, MD), and other DAT blockers are not recognized as substrates even though they can access the primary substrate binding pocket. Proteins 2010. (c) 2009 Wiley-Liss, Inc.

Benchmarking docking and scoring protocol for the identification of potential acetylcholinesterase inhibitors

Abstract: Acetylcholinesterase (AChE) plays a crucial role in nerve impulse transmission at cholinergic synapses by rapid hydrolysis of the neurotransmitter acetylcholine (ACh). AChE has become an important drug target because partial inhibition of AChE results in modest increase in ACh levels that can have therapeutic benefits, thus AChE inhibitors have proved useful in the symptomatic treatment of Alzheimer's disease. To establish an effective docking protocol for virtual screening of AChE, a comparative molecular docking study was performed. For this purpose six docking/scoring approaches (AutoDock, FlexX, MOE, Surflex-Dock, GOLD and FRED) were compared to determine their ability to reproduce the binding poses in twenty six complexes of AChE. Docking accuracy was evaluated by calculating the RMSD of the docked complexes. FRED was found to be the best in reproducing the experimental pose by placing it near the top of its ranking. The performance of scoring functions was evaluated by identifying known actives out of large database of inactive compounds. A dataset of 5000 "drug like" decoys were retrieved from NCI database and docked into the binding site of AChE with six known inhibitors using FRED in combination with five scoring functions, i.e., Chemgauss2, Chemgauss3, ChemScore, Shapegauss and PLP. The poses obtained by FRED were re-scored using GOLD score, ChemScore and ASP as implemented in GOLD while G_Score, D_Score, ChemScore and PMF as implemented in the CScore module of SYBYL7.3. D_Score presented significantly better enrichment than others and 50% of the active inhibitors were identified in top 20% of the ranked database.

Receptor-Based Discovery of a Plasmalemmal Monoamine Transporter Inhibitor via High Throughput Docking and Pharmacophore Modeling.

Abstract: Recognition of psychostimulants such as cocaine and the amphetamines by the dopamine transporter (DAT) protein is principally responsible for the euphoria and addiction associated with these drugs. Using as a template the crystal structure of a distantly related bacterial leucine transporter, 3-D DAT computer molecular models have been generated. Ligand docking to such models has revealed potential substrate and inhibitor binding pockets, subsequently confirmed by in vitro pharmacology. An inhibitor pocket defined by the DAT model to be within the "extracellular vestibule", just to the extracellular side of the external gate of the primary substrate pocket, was used for virtual screening of a structural library of compounds. High-throughput docking and application of pharmacophore constraints within this vestibular inhibitor pocket identified a compound structurally dissimilar to the classic monoamine (dopamine, norepinephrine and serotonin) transporter (MAT) inhibitors. The compound displaced binding of radiolabeled cocaine analogs at all three MATs, usually with nanomolar K(i) values and within two fold of cocaine's affinity at the norepinephrine transporter. Although a very weak dopamine uptake inhibitor itself, this compound reduced by three fold the potency of cocaine in inhibiting DAT-mediated cellular uptake of dopamine. To our knowledge, the present findings are the first to successfully employ "receptor-based" computer modeling to identify moderate-to-high affinity MAT ligands. In silico ligand screening using MAT models provides a rapid, low cost discovery process that should accelerate identification of novel ligand scaffolds and provide lead compounds in combating psychostimulant addiction and in treating other monoamine-related CNS diseases.

Circular Dichroism and UV Resonance Raman Study of the Impact of Salts and Alcohols on the Gibbs Free Energy Landscape of an α‐helical Peptide

Abstract: We used CD and UV resonance Raman spectroscopy to study the impact of alcohols on the conformational equilibria and relative Gibbs free energy landscapes along the Ramachandran Ψ-coordinate of a mainly poly-Ala peptide, AP with an AAAAA(AAARA)3A sequence. 2,2,2-Trifluoroethanol (TFE) most stabilizes the α-helix-like conformations, followed by ethanol, methanol, and pure water. The π-bulge conformation is stabilized more than the α-helix, while the 310-helix is destabilized due to the alcohol-increased hydrophobicity. Turns are also stabilized by alcohols. We also found that while TFE induces more α-helices, it favors multiple, shorter helix segments.