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Lipid nanoemulsions are promising nanomaterials for drug delivery applications in food, pharmaceutical and cosmetic industries. Despite the noteworthy commercial interest, little is known about their supramolecular organization, especially about how such multicomponent formulations interact with cell membranes. In the present work, coarse-grained molecular dynamics simulations have been employed to study the self-assembly of a 15-component lipid nanoemulsion droplet containing vitamins A and E for skin delivery. Our results display aspects of the unique "onion-like" agglomeration between the chemical constituents in the different layers of the lipid nanodroplet. Vitamin E molecules are more concentrated in the center of the droplet together with other hydrophobic constituents such as the triglycerides with long tails. On the other hand, vitamin A occupies an intermediate layer between the core and the co-emulsifier surface of the nanodroplet, together with lecithin phospholipids. Coarse-grained molecular dynamics simulations were also performed to provide insight into the first steps involved in absorption and penetration of the nanodroplet through skin membrane models, representing an intracellular (hair follicle infundibulum) and intercellular pathway (stratum corneum) through the skin. Our data provide a first view on the complex organization of commercial nanoemulsion and its interaction with skin membranes. We expect our results to open the way towards the rational design of such nanomaterials.

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A technical overview of the High Performance Collision Cross Section (HPCCS) software for accurate and efficient calculations of collision cross sections for molecular ions ranging from small organic molecules to large protein complexes is presented. The program uses helium or nitrogen as buffer gas with considerable gains in computer time compared to publicly available codes under the Trajectory Method approximation. HPCCS is freely available under the Academic Use License at https://github.com/cepid-cces/hpccs .

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Since the commercial introduction of Ion Mobility coupled with Mass Spectrometry (IM-MS) devices in 2003, a large number of research laboratories have embraced the technique. IM-MS is a fairly rapid experiment used as a molecular separation tool and to obtain structural information. The interpretation of IM-MS data is still challenging and relies heavily on theoretical calculations of the molecule's collision cross section (CCS) against a buffer gas. Here, a new software (HPCCS) is presented, which performs CCS calculations using high perfomance computing techniques. Based on the trajectory method, HPCCS can accurately calculate CCS for a great variety of molecules, ranging from small organic molecules to large protein complexes, using helium or nitrogen as buffer gas with considerable gains in computer time compared to publicly available codes under the same level of theory. HPCCS is available as free software under the Academic Use License at https://github.com/cepid-cces/hpccs. © 2018 Wiley Periodicals, Inc.

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The natural ligand 17ß-estradiol (E2) is so far believed to induce a unique agonist-bound active conformation in the ligand binding domain (LBD) of the estrogen receptors (ERs). Both subtypes, ERα and ERß, are transcriptionally activated in the presence of E2 with ERß being somewhat less active than ERα under similar conditions. The molecular bases for this intriguing behavior are mainly attributed to subtype differences in the amino-terminal domain of these receptors. However, structural details that confer differences in the molecular response of ER LBDs to E2 still remain elusive. In this study, we present a new crystallographic structure of the ERß LBD bound to E2 in which H12 assumes an alternative conformation that resembles antagonist ERs structures. Structural observations and molecular dynamics simulations jointly provide evidence that alternative ERß H12 position could correspond to a stable conformation of the receptor under physiological pH conditions. Our findings shed light on the unexpected role of LBD in the lower functional response of ERß subtype.

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The peroxisome proliferator-activated receptor γ (PPARγ) ligands are important therapeutic drugs for the treatment of type 2 diabetes, obesity and cardiovascular diseases. In particular, partial agonists and non-agonists are interesting targets to reduce glucose levels, presenting few side effects in comparison to full agonists. In this work, we present a set of CHARMM-based parameters of a molecular mechanics force field for two PPARγ ligands, GQ16 and SR1664. GQ16 belongs to the thiazolidinedione class of drugs and it is a PPARγ partial agonist that has been shown to promote the "browning" of white adipose tissue. SR1664 is the precursor of the PPARγ non-agonist class of ligands that activates PPARγ in a non-classical manner. Here, we use quantum chemical calculations consistent with the CHARMM protocol to obtain bonded and non-bonded parameters, including partial atomic charges and effective torsion potentials for both molecules. The newly parameterized models were evaluated by examining the behavior of GQ16 and SR1664 free in water and bound to the ligand binding pocket of PPARγ using molecular dynamics simulations. The potential parameters derived here are readily transferable to a variety of pharmaceutical compounds and similar PPARγ ligands.

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The peroxisome proliferator-activated receptor γ (PPARγ) is an important transcription factor that plays a major role in the regulation of glucose and lipid metabolisms and has, therefore, many implications in modern-life metabolic disorders such as diabetes, obesity, and cardiovascular diseases. Phosphorylation of PPARγ by the cyclin-dependent kinase 5 (Cdk5) has been recently proved to promote obesity and loss of insulin sensitivity. The inhibition of this reaction is currently being pursued to develop PPARγ ligands for type 2 diabetes treatments. The knowledge of the protein-protein interactions between Cdk5/p25 and PPARγ can be an important asset for better understanding of the molecular basis of the Cdk5-meditated phosphorylation of PPARγ and its inhibition. By means of a computational approach that combines protein-protein docking and adaptive biasing force molecular dynamics simulations, we obtained PPARγ-Cdk5/p25 structural models that are consistent with the mechanism of the enzymatic reaction and with overall structural features of the full length PPARγ-RXRα heterodimer bound to DNA. In addition to the active site, our model shows that the interacting regions between the two proteins should involve two distal docking sites, comprising the PPARγ Ω-loop and Cdk5 N-terminal lobe and the PPARγ ß-sheet and Cdk5 C-terminal lobe. These sites are related to PPARγ transactivation and directly interact with PPARγ ligands. Our results suggest that ß-sheets and Ω-loop stabilization promoted by PPARγ agonists could be important to inhibit Cdk5-mediated phosphorylation.

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Peroxisome proliferator-activated receptors (PPARs) are members of a superfamily of nuclear transcription factors. They are involved in mediating numerous physiological effects in humans, including glucose and lipid metabolism. PPARα ligands effectively treat dyslipidemia and have significant antiinflammatory and anti-atherosclerotic activities. These effects and their ligand-dependent activity make nuclear receptors obvious targets for drug design. Here, we present the structure of the human PPARα in complex with WY14643, a member of fibrate class of drug, and a widely used PPAR activator. The crystal structure of this complex suggests that WY14643 induces activation of PPARα in an unusual bipartite mechanism involving conventional direct helix 12 stabilization and an alternative mode that involves a second ligand in the pocket. We present structural observations, molecular dynamics and activity assays that support the importance of the second site in WY14643 action. The unique binding mode of WY14643 reveals a new pattern of nuclear receptor ligand recognition and suggests a novel basis for ligand design, offering clues for improving the binding affinity and selectivity of ligand. We show that binding of WY14643 to PPARα was associated with antiinflammatory disease in a human corneal cell model, suggesting possible applications for PPARα ligands.

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Thiazolidinediones (TZDs) act through peroxisome proliferator activated receptor (PPAR) γ to increase insulin sensitivity in type 2 diabetes (T2DM), but deleterious effects of these ligands mean that selective modulators with improved clinical profiles are needed. We obtained a crystal structure of PPARγ ligand binding domain (LBD) and found that the ligand binding pocket (LBP) is occupied by bacterial medium chain fatty acids (MCFAs). We verified that MCFAs (C8-C10) bind the PPARγ LBD in vitro and showed that they are low-potency partial agonists that display assay-specific actions relative to TZDs; they act as very weak partial agonists in transfections with PPARγ LBD, stronger partial agonists with full length PPARγ and exhibit full blockade of PPARγ phosphorylation by cyclin-dependent kinase 5 (cdk5), linked to reversal of adipose tissue insulin resistance. MCFAs that bind PPARγ also antagonize TZD-dependent adipogenesis in vitro. X-ray structure B-factor analysis and molecular dynamics (MD) simulations suggest that MCFAs weakly stabilize C-terminal activation helix (H) 12 relative to TZDs and this effect is highly dependent on chain length. By contrast, MCFAs preferentially stabilize the H2-H3/ß-sheet region and the helix (H) 11-H12 loop relative to TZDs and we propose that MCFA assay-specific actions are linked to their unique binding mode and suggest that it may be possible to identify selective PPARγ modulators with useful clinical profiles among natural products.

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Nuclear hormone receptors (NRs) form a family of transcription factors that mediate cellular responses initiated by hormone binding. It is generally recognized that the structure and dynamics of the C-terminal helix 12 (H12) of NRs' ligand binding domain (LBD) are fundamental to the recognition of coactivators and corepressors that modulate receptor function. Here we study the role of three mutations in the I280 residue of H12 of thyroid hormone receptors using site-directed mutagenesis, functional assays, and molecular dynamics simulations. Although residues at position 280 do not interact with coactivators or with the ligand, we show that its mutations can selectively block coactivator and corepressor binding, and affect hormone binding affinity differently. Molecular dynamics simulations suggest that ligand affinity is reduced by indirectly displacing the ligand in the binding pocket, facilitating water penetration and ligand destabilization. Mutations I280R and I280K link H12 to the LBD by forming salt bridges with E457 in H12, stabilizing H12 in a conformation that blocks both corepressor and coactivator recruitment. The I280M mutation, in turn, blocks corepressor binding, but appears to enhance coactivator affinity, suggesting stabilization of H12 in agonist conformation.

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The ligand binding domain (LBD) of nuclear hormone receptors adopts a very compact, mostly alpha-helical structure that binds specific ligands with very high affinity. We use circular dichroism spectroscopy and high-temperature molecular dynamics simulations to investigate unfolding of the LBDs of thyroid hormone receptors (TRs). A molecular description of the denaturation mechanisms is obtained by molecular dynamics simulations of the TRalpha and TRbeta LBDs in the absence and in the presence of the natural ligand Triac. The simulations show that the thermal unfolding of the LBD starts with the loss of native contacts and secondary structure elements, while the structure remains essentially compact, resembling a molten globule state. This differs from most protein denaturation simulations reported to date and suggests that the folding mechanism may start with the hydrophobic collapse of the TR LBDs. Our results reveal that the stabilities of the LBDs of the TRalpha and TRbeta subtypes are affected to different degrees by the binding of the isoform selective ligand Triac and that ligand binding confers protection against thermal denaturation and unfolding in a subtype specific manner. Our simulations indicate two mechanisms by which the ligand stabilizes the LBD: (1) by enhancing the interactions between H8 and H11, and the interaction of the region between H1 and the Omega-loop with the core of the LBD, and (2) by shielding the hydrophobic H6 from hydration.

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