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Sensing hydration and behavior of pyrene in POPC and POPC/cholesterol bilayers: A molecular dynamics study

Publication . Loura, Luís M. S.; Martins do Canto, Antonio M. T.; Martins, Jorge

Molecular dynamics (MD) simulations of bilayers of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) with varying amounts of cholesterol (0, 5, 20, and 40 mol%) were carried out in the absence and presence of inserted pyrene molecules. Both fluorophore and bilayer parameters were computed, for characterization of probe location and dynamics, as well as its effects on the host bilayer. In agreement with previous studies in fluid disordered bilayers, pyrene prefers to be located in the hydrophobic acyl chain region of POPC bilayers, close to the glycerol group of lipid molecules and causes ordering of the lipid acyl chains. However, incorporation of pyrene in binary POPC/cholesterol bilayers decreases the acyl chain order parameter (especially near the end of the chains), opposing the ordering effect of cholesterol. These effects are modest and mainly felt locally. Significantly, as the bilayer is enriched with cholesterol, the relative position of pyrene and the POPC carbonyl and phosphocholine groups is invariant, and the local water density around the probe decreases. This work clarifies and supports the cautious use of pyrene Ham effect to effectively measure equivalent polarity in lipid bilayers. Within the time scale of the MD simulations, which is of the magnitude of the fluorescence lifetime of pyrene, the thermally averaged polarity of lipid bilayers is nearly out of influence of spurious uncertainty in the transverse location of pyrene in the bilayers. This renders the values of equivalent polarity measurements through the pyrene Ham effect more reliable and reproducible than previously expected. (C) 2012 Elsevier B.V. All rights reserved.

2013-03Journal article

Open Access

Behavior of pyrene as a polarity probe in palmitoylsphingomyelin and palmitoylsphingomyelin/cholesterol bilayers: A molecular dynamics simulation study

Publication . do Canto, António M. T. M.; Santos, Patrícia D.; Martins, Jorge; Loura, Luís M. S.

Pyrene is a polycyclic aromatic hydrocarbon noted for its remarkable optical spectroscopic properties. Among its uses as a fluorescent probe, measurement of lipid bilayer's equivalent polarity through the pyrene Ham effect stands out. To this effect, the ratio of the intensities of the first and third vibronic bands (I-1/I-3) in its emission spectrum of pyrene is measured. However, issues concerning the precise location of bilayer-inserted pyrene and the possibility of probe-induced perturbation of host bilayer properties are potential sources of concern in this regard. Atomistic molecular dynamics simulations constitute a useful method for the characterization of lipid membrane systems, and, in particular, to understand the behavior of fluorescence probes upon incorporation in lipid bilayers. In this report, we present a detailed characterization of the behavior of pyrene in fluid N-palmitoylsphingomyelin (PSM) and PSM/cholesterol membranes, with emphasis on the degree of proximity between the probe and water molecules inside bilayers, related to the use of pyrene to measure equivalent lipid bilayer polarity. It is concluded that pyrene exerts minor effects on bilayer properties, with slight local disordering being apparent for high cholesterol content. Whereas rotation and lateral diffusion of pyrene are greatly slowed in cholesterol rich systems, its relative transverse location is not significantly affected. While hydration of PSM bilayers, as sensed by pyrene, is already low compared to that of fluid phosphatidylcholine, it becomes even smaller for high cholesterol mole fraction at the studied temperature. (C) 2014 Elsevier B.V. All rights reserved.

2015-09Journal article

Open Access

Ion pumps as biological targets for decavanadate

Publication . Aureliano, M.

The putative applications of poly-, oligo- and mono-oxometalates in biochemistry, biology, pharmacology and medicine are rapidly attracting interest. In particular, these compounds may act as potent ion pump inhibitors and have the potential to play a role in the treatment of e.g. ulcers, cancer and ischemic heart disease. However, the mechanism of action is not completely understood in most cases, and even remains largely unknown in other cases. In the present review we discuss the most recent insights into the interaction between mono- and polyoxometalate ions with ion pumps, with a particular focus on the interaction of decavanadate with Ca2+- ATPase. We also compare the proposed mode of action with those of established ion pump inhibitors which are currently in therapeutic use. Of the 18 classes of compounds which are known to act as ion pump inhibitors, the complete mechanism of inhibition is only known for a handful. It has, however, been established that most ion pump inhibitors bind mainly to the E2 ion pump conformation within the membrane domain from the extracellular side and block the cation release. Polyoxometalates such as decavanadate, in contrast, interact with Ca2+- ATPase near the nucleotide binding site domain or at a pocket involving several cytoplasmic domains, and therefore needs to cross through the membrane bilayer. In contrast to monomeric vanadate, which only binds to the E2 conformation, decavanadate binds to all protein conformations, i.e. E1, E1P, E2 and E2P. Moreover, the specific interaction of decavanadate with sarcoplasmic reticulum Ca2+- ATPase has been shown to be non-competitive with respect to ATP and induces protein cysteine oxidation with concomitant vanadium reduction which might explain the high inhibitory capacity of V10, (IC50=15 µM) which is quite similar to the majority of the established therapeutic drugs.

2013Journal article

Open Access

Synthesis and structure of 2-substituted pyrene-derived scaffolds

Publication . Cabral, Lília; Henriques, Marta Sofia; Paixão, José António; Lurdes S. Cristiano, M.

Pyrenes bear a propensity to form fluorescent excimers, and thus this chromophore is often found in sensors and fluorescent probes. 2-FunctionaIized pyrenes are of particular interest, however the preparation of these scaffolds is not trivial, involving synthetic routes that require 4,5,9,10-tetrahydropyrene as a key intermediate. Herein, the development and optimization of routes for the synthesis of 2-functionalized pyrene-derived building blocks, with potential to be used as tags in the preparation of fluorescent probes, is described. Additionally, the crystal structures of ethyl 4,5,9,10-tetrahydro-2-pyrene-5-oxopentanoate and 2-acetyl-4,5,9,10-tetrahydropyrene revealed distinct conformations of the saturated tetrahydropyrene rings. (C) 2017 Elsevier Ltd. All rights reserved.

2017-11Journal article

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