Automated Builder and Database of Protein/Membrane Complexes for Molecular Dynamics Simulations

Jo, Sunhwan; Kim, Tae-Hoon; Im, Wonpil

doi:10.1371/journal.pone.0000880

Cited by 1,075 publications

(967 citation statements)

References 36 publications

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“…Initial peptide structures were built with CHARMM 55,56 in extended conformations. The phospholipid bilayers with a single copy of peptide in aqueous phase were created by using CHARMM-GUI [57][58][59] and all molecular dynamics simulations were conducted with GROMACS 4.5.4 or 4.5.6. 60 The bilayer systems contained the following molecules: 50 (2 × 25) 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) molecules, one peptide, eight chloride, and eight sodium ions and TIP3P water.…”

Section: Computationalmentioning

confidence: 99%

Permeation of the three aromatic dipeptides through lipid bilayers: Experimental and computational study

Lee

Kuczera

Middaugh

et al. 2016

The Journal of Chemical Physics

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The time-resolved parallel artificial membrane permeability assay with fluorescence detection and comprehensive computer simulations are used to study the passive permeation of three aromatic dipeptides—N-acetyl-phenylalanineamide (NAFA), N-acetyltyrosineamide (NAYA), and N-acetyl-tryptophanamide (NATA) through a 1,2-dioleoyl-sn-glycero-3-phospocholine (DOPC) lipid bilayer. Measured permeation times and permeability coefficients show fastest translocation for NAFA, slowest for NAYA, and intermediate for NATA under physiological temperature and pH. Computationally, we perform umbrella sampling simulations to model the structure, dynamics, and interactions of the peptides as a function of z, the distance from lipid bilayer. The calculated profiles of the potential of mean force show two strong effects—preferential binding of each of the three peptides to the lipid interface and large free energy barriers in the membrane center. We use several approaches to calculate the position-dependent translational diffusion coefficients D(z), including one based on numerical solution the Smoluchowski equation. Surprisingly, computed D(z) values change very little with reaction coordinate and are also quite similar for the three peptides studied. In contrast, calculated values of sidechain rotational correlation times τrot(z) show extremely large changes with peptide membrane insertion—values become 100 times larger in the headgroup region and 10 times larger at interface and in membrane center, relative to solution. The peptides’ conformational freedom becomes systematically more restricted as they enter the membrane, sampling α and β and C7eq basins in solution, α and C7eq at the interface, and C7eq only in the center. Residual waters of solvation remain around the peptides even in the membrane center. Overall, our study provides an improved microscopic understanding of passive peptide permeation through membranes, especially on the sensitivity of rotational diffusion to position relative to the bilayer.

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Section: Computationalmentioning

confidence: 99%

Permeation of the three aromatic dipeptides through lipid bilayers: Experimental and computational study

Lee

Kuczera

Middaugh

et al. 2016

The Journal of Chemical Physics

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“…The trajectories described therein form the basis for the current analysis. Briefly, four independent all-atom models of monomeric vSGLT based on the inward-occluded, galactose-bound state X-ray structure [Protein Data Bank (PDB) ID code 3DH4] (12), were embedded in a POPE lipid bilayer, using MODELLER (30) and the CHARMM-GUI Membrane Builder (31). The models were parameterized using the CHARMM22 force field (32) with CMAP corrections (33) for the protein, the CHARMM36 lipid force field (34), the CHARMM pyranose monosaccharides parameter set for the galactose (35), and the TIP3P explicit water model (36).…”

Section: Methodsmentioning

confidence: 99%

Stochastic steps in secondary active sugar transport

Adelman

Ghezzi

Bisignano

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Secondary active transporters, such as those that adopt the leucinetransporter fold, are found in all domains of life, and they have the unique capability of harnessing the energy stored in ion gradients to accumulate small molecules essential for life as well as expel toxic and harmful compounds. How these proteins couple ion binding and transport to the concomitant flow of substrates is a fundamental structural and biophysical question that is beginning to be answered at the atomistic level with the advent of high-resolution structures of transporters in different structural states. Nonetheless, the dynamic character of the transporters, such as ion/substrate binding order and how binding triggers conformational change, is not revealed from static structures, yet it is critical to understanding their function. Here, we report a series of molecular simulations carried out on the sugar transporter vSGLT that lend insight into how substrate and ions are released from the inward-facing state of the transporter. Our simulations reveal that the order of release is stochastic. Functional experiments were designed to test this prediction on the human homolog, hSGLT1, and we also found that cytoplasmic release is not ordered, but we confirmed that substrate and ion binding from the extracellular space is ordered. Our findings unify conflicting published results concerning cytoplasmic release of ions and substrate and hint at the possibility that other transporters in the superfamily may lack coordination between ions and substrate in the inward-facing state.T ransport of sugar molecules across membranes is virtually ubiquitous in biology, and it is of central importance in human health. The uptake of glucose is especially crucial due to its pivotal role in cellular metabolism and energy production. In mammals, glucose is absorbed in the small intestine and kidney via sodium-dependent glucose transporters (SGLTs), which localize to the apical membrane and concentrate glucose in the epithelia. SGLTs fall into the large leucine-transporter (LeuT) structural family of secondary active transporters that have evolved to concentrate a wide array of substrates across membranes using the energy stored in the Na + electrochemical potential gradient. For symporters in this family, transport occurs by an alternating access mechanism (1) in which the transporter first binds ligands in an outward-facing conformation, and then transitions to an inwardfacing conformation that releases the cargo to the cytoplasm. The order of ion and substrate binding and unbinding is likely tied to the function of the transporter, making it possible to convert the energy stored in the ion gradient into a substrate gradient, and vice versa when these proteins operate in reverse.Extensive biochemical uptake assays and electrophysiological studies of SGLTs have led to a view of Na + /glucose cotransport in which Na + binding precedes sugar binding on the external face of the transporter. Kinetic models adhering to this mechanism satisfactorily account fo...

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“…Results were validated for WT NIS with explicit water and a POPC membrane using GROMACS 5.0 (72,73). The initial water molecules and the bilayer were built using the web-based automated builder CHARMM-GUI (74)(75)(76). The protein model used was the same as that used in the implicit solvent simulation.…”

Section: Methodsmentioning

confidence: 99%

“…The equilibrated system was extended into a 100-ns-long MD production simulation using a Nose-Hoover thermostat. The LINear Constant Solvent (LINCS) algorithm was used to constrain bonds, and the Particle Mesh Ewald algorithm was used to treat long-range electrostatic interactions during equilibration of the system and MD production runs (72)(73)(74)(75)(76)(77).…”

Section: Methodsmentioning

confidence: 99%

Na ⁺ coordination at the Na2 site of the Na ⁺ /I ⁻ symporter

Ferrandino

Nicola

Sánchez

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The sodium/iodide symporter (NIS) mediates active I− transport in the thyroid—the first step in thyroid hormone biosynthesis—with a 2 Na+: 1 I− stoichiometry. The two Na+ binding sites (Na1 and Na2) and the I− binding site interact allosterically: when Na+ binds to a Na+ site, the affinity of NIS for the other Na+ and for I− increases significantly. In all Na+-dependent transporters with the same fold as NIS, the side chains of two residues, S353 and T354 (NIS numbering), were identified as the Na+ ligands at Na2. To understand the cooperativity between the substrates, we investigated the coordination at the Na2 site. We determined that four other residues—S66, D191, Q194, and Q263—are also involved in Na+ coordination at this site. Experiments in whole cells demonstrated that these four residues participate in transport by NIS: mutations at these positions result in proteins that, although expressed at the plasma membrane, transport little or no I−. These residues are conserved throughout the entire SLC5 family, to which NIS belongs, suggesting that they serve a similar function in the other transporters. Our findings also suggest that the increase in affinity that each site displays when an ion binds to another site may result from changes in the dynamics of the transporter. These mechanistic insights deepen our understanding not only of NIS but also of other transporters, including many that, like NIS, are of great medical relevance.

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Automated Builder and Database of Protein/Membrane Complexes for Molecular Dynamics Simulations

Cited by 1,075 publications

References 36 publications

Permeation of the three aromatic dipeptides through lipid bilayers: Experimental and computational study

Permeation of the three aromatic dipeptides through lipid bilayers: Experimental and computational study

Stochastic steps in secondary active sugar transport

Na ⁺ coordination at the Na2 site of the Na ⁺ /I ⁻ symporter

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Automated Builder and Database of Protein/Membrane Complexes for Molecular Dynamics Simulations

Cited by 1,075 publications

References 36 publications

Permeation of the three aromatic dipeptides through lipid bilayers: Experimental and computational study

Permeation of the three aromatic dipeptides through lipid bilayers: Experimental and computational study

Stochastic steps in secondary active sugar transport

Na + coordination at the Na2 site of the Na + /I − symporter

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Na ⁺ coordination at the Na2 site of the Na ⁺ /I ⁻ symporter