This review focuses on the influence of oxidized phosphatidylcholines (oxPCs) on the biophysical properties of model membranes and is limited to fluorescence, EPR, and MD studies. OxPCs are divided into two classes: A) hydroxy- or hydroperoxy-dieonyl phospatidylcholines, B) phospatidylcholines with oxidized and truncated chains with either aldehyde or carboxylic group. It was shown that the presence of the investigated oxPCs in phospholipid model membranes may have the following consequences: 1) decrease of the lipid order, 2) lowering of phase transition temperatures, 3) lateral expansion and thinning of the bilayer, 4) alterations of bilayer hydration profiles, 5) increased lipid mobility, 6) augmented flip-flop, 7) influence on the lateral phase organisation, and 8) promotion of water defects and, under extreme conditions (i.e. high concentrations of class B oxPCs), disintegration of the bilayer. The effects of class A oxPCs appear to be more moderate than those observed or predicted for class B. Many of the abovementioned findings are related to the ability of the oxidized chains of certain oxPCs to reorient toward the water phase. Some of the effects appear to be moderated by the presence of cholesterol. Although those biophysical alternations are found at oxPC concentrations higher than the total oxPC concentrations found under physiological conditions, certain organelles may reach such elevated oxPC concentrations locally. It is a challenge for the future to correlate the biophysics of oxidized phospholipids to metabolic studies in order to define the significance of the findings presented herein for pathophysiology. This article is part of a Special Issue entitled: Oxidized phospholipids-their properties and interactions with proteins.

• MeSH
• biofyzikální jevy * MeSH
• elektronová paramagnetická rezonance MeSH
• fluorescence MeSH
• fosfolipidy chemie   metabolismus   MeSH
• lidé MeSH
• lipidové dvojvrstvy chemie   metabolismus   MeSH
• oxidace-redukce MeSH
• simulace molekulární dynamiky * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• fosfolipidy MeSH
• lipidové dvojvrstvy MeSH

• Klíčová slova
• caveolae, cell membrane, fluorescence microscopy, membrane properties, membrane trafficking, nanodomains, superresolution microscopy, tetraspanins,
• Publikační typ
• úvodníky MeSH

The ability of the four-stranded guanine (G)-DNA motif to incorporate nonstandard guanine analogue bases 6-oxopurine (inosine, I), 6-thioguanine (tG), and 6-thiopurine (tI) has been investigated using large-scale molecular dynamics simulations. The simulations suggest that a G-DNA stem can incorporate inosines without any marked effect on its structure and dynamics. The all-inosine quadruplex stem d(IIII)(4) shows identical dynamical properties as d(GGGG)(4) on the nanosecond time scale, with both molecular assemblies being stabilized by monovalent cations residing in the channel of the stem. However, simulations carried out in the absence of these cations show dramatic differences in the behavior of d(GGGG)(4) and d(IIII)(4). Whereas vacant d(GGGG)(4) shows large fluctuations but does not disintegrate, vacant d(IIII)(4) is completely disrupted within the first nanosecond. This is a consequence of the lack of the H-bonds involving the N2 amino group that is not present in inosine. This indicates that formation of the inosine quadruplex could involve entirely different intermediate structures than formation of the guanosine quadruplex, and early association of cations in this process appears to be inevitable. In the simulations, the incorporation of 6-thioguanine and 6-thiopurine sharply destabilizes four-stranded G-DNA structures, in close agreement with experimental data. The main reason is the size of the thiogroup leading to considerable steric conflicts and expelling the cations out of the channel of the quadruplex stem. The G-DNA stem can accommodate a single thioguanine base with minor perturbations. Incorporation of a thioguanine quartet layer is associated with a large destabilization of the G-DNA stem whereas the all-thioguanine quadruplex immediately collapses.

• MeSH
• biofyzika MeSH
• biofyzikální jevy MeSH
• DNA chemie   MeSH
• inosin chemie   MeSH
• iontové kanály chemie   MeSH
• konformace nukleové kyseliny * MeSH
• merkaptopurin chemie   MeSH
• molekulární modely MeSH
• sodík chemie   MeSH
• termodynamika MeSH
• thioguanin chemie   MeSH
• vodíková vazba MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA MeSH
• inosin MeSH
• iontové kanály MeSH
• merkaptopurin MeSH
• sodík MeSH
• thioguanin MeSH

Variegate porphyria is caused by mutations in the protoporphyrinogen oxidase IX (PPOX, EC 1.3.3.4) gene, resulting in reduced overall enzymatic activity of PPOX in human tissues. Recently, we have identified the His333Arg mutation in the PPOX protein (PPOX(H333R)) as a putative founder mutation in the Moroccan Jewish population. Herein we report the molecular characterization of PPOX(H333R) in vitro and in cells. Purified recombinant PPOX(H333R) did not show any appreciable enzymatic activity in vitro, corroborating the clinical findings. Biophysical experiments and molecular modeling revealed that PPOX(H333R) is not folded properly and fails to adopt its native functional three-dimensional conformation due to steric clashes in the vicinity of the active site of the enzyme. On the other hand, PPOX(H333R) subcellular distribution, as evaluated by live-cell confocal microscopy, is unimpaired suggesting that the functional three-dimensional fold is not required for efficient transport of the polypeptide chain into mitochondria. Overall, the data presented here provide molecular underpinnings of the pathogenicity of PPOX(H333R) and might serve as a blueprint for deciphering whether a given PPOX variant represents a disease-causing mutation.

• Klíčová slova
• Heme synthesis, Protoporphyrinogen oxidase IX, Variegate porphyria,
• MeSH
• biofyzikální jevy MeSH
• buněčné linie MeSH
• flavoproteiny chemie   genetika   izolace a purifikace   MeSH
• kinetika MeSH
• lidé MeSH
• mitochondriální proteiny chemie   genetika   izolace a purifikace   MeSH
• molekulární modely MeSH
• multimerizace proteinu MeSH
• mutace genetika   MeSH
• protoporfyrinogenoxidasa chemie   genetika   izolace a purifikace   MeSH
• sekvence aminokyselin MeSH
• stabilita enzymů MeSH
• subcelulární frakce metabolismus   MeSH
• teplota MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• flavoproteiny MeSH
• mitochondriální proteiny MeSH
• PPOX protein, human MeSH Prohlížeč
• protoporfyrinogenoxidasa MeSH

• MeSH
• biofyzika MeSH
• biofyzikální jevy MeSH
• DNA chemie   MeSH
• fosfáty chemie   MeSH
• konformace nukleové kyseliny MeSH
• nukleové kyseliny chemie   MeSH
• oligodeoxyribonukleotidy chemie   MeSH
• oligonukleotidy chemie   MeSH
• organofosfonáty chemie   MeSH
• sekvence nukleotidů MeSH
• termodynamika MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA MeSH
• fosfáty MeSH
• nukleové kyseliny MeSH
• oligodeoxyribonukleotidy MeSH
• oligonukleotidy MeSH
• organofosfonáty MeSH

MOTIVATION: Metallothionein-III (MT-III) displays neuro-inhibitory activity and is involved in the repair of neuronal damage. An altered expression level of MT-III suggests that it could be a mitigating factor in Alzheimer's disease (AD) neuronal dysfunction. Currently there are limited marketed drugs available against MT-III. The inhibitors are mostly pseudo-peptide based with limited ADMET. In our present study, available database InterBioScreen (natural compounds) was screened out for MT-III. Pharmacodynamics and pharmacokinetic studies were performed. Molecular docking and simulations of top hit molecules were performed to study complex stability. RESULTS: Study reveals potent selective molecules that interact and form hydrogen bonds with amino acids Ser-6 and Lys-22 are common to established melatonin inhibitors for MT-III. These include DMHMIO, MCA B and s27533 derivatives. The ADMET profiling was better with comparable interaction energy values. It includes properties like blood brain barrier, hepatotoxicity, druggability, mutagenicity and carcinogenicity. Molecular dynamics studies were performed to validate our findings.

• Klíčová slova
• ADMET, Alzheimer’s disease, Metallothionein-III, Molecular dynamics, Virtual screening,
• MeSH
• Alzheimerova nemoc metabolismus   patologie   MeSH
• biofyzikální jevy MeSH
• lidé MeSH
• metalothionein 3 MeSH
• proteiny nervové tkáně antagonisté a inhibitory   chemie   MeSH
• simulace molekulární dynamiky MeSH
• simulace molekulového dockingu MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• metalothionein 3 MeSH
• proteiny nervové tkáně MeSH

Biophysical properties of the tear film lipid layer are studied at the molecular level employing coarse grain molecular dynamics (MD) simulations with a realistic model of the human tear film. In this model, polar lipids are chosen to reflect the current knowledge on the lipidome of the tear film whereas typical Meibomian-origin lipids are included in the thick non-polar lipids subphase. Simulation conditions mimic those experienced by the real human tear film during blinks. Namely, thermodynamic equilibrium simulations at different lateral compressions are performed to model varying surface pressure, and the dynamics of the system during a blink is studied by non-equilibrium MD simulations. Polar lipids separate their non-polar counterparts from water by forming a monomolecular layer whereas the non-polar molecules establish a thick outermost lipid layer. Under lateral compression, the polar layer undulates and a sorting of polar lipids occurs. Moreover, formation of three-dimensional aggregates of polar lipids in both non-polar and water subphases is observed. We suggest that these three-dimensional structures are abundant under dynamic conditions caused by the action of eye lids and that they act as reservoirs of polar lipids, thus increasing stability of the tear film.

• MeSH
• biofyzika * MeSH
• lidé MeSH
• lipidy chemie   MeSH
• slzy chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• lipidy MeSH

Skin plays an important role in protection, metabolism, thermoregulation, sensation, and excretion whilst being consistently exposed to environmental aggression, including biotic and abiotic stresses. During the generation of oxidative stress in the skin, the epidermal and dermal cells are generally regarded as the most affected regions. The participation of reactive oxygen species (ROS) as a result of environmental fluctuations has been experimentally proven by several researchers and is well known to contribute to ultra-weak photon emission via the oxidation of biomolecules (lipids, proteins, and nucleic acids). More recently, ultra-weak photon emission detection techniques have been introduced to investigate the conditions of oxidative stress in various living systems in in vivo, ex vivo and in vitro studies. Research into two-dimensional photon imaging is drawing growing attention because of its application as a non-invasive tool. We monitored spontaneous and stress-induced ultra-weak photon emission under the exogenous application of a Fenton reagent. The results showed a marked difference in the ultra-weak photon emission. Overall, these results suggest that triplet carbonyl (3C=O∗) and singlet oxygen (1O2) are the final emitters. Furthermore, the formation of oxidatively modified protein adducts and protein carbonyl formation upon treatment with hydrogen peroxide (H2O2) were observed using an immunoblotting assay. The results from this study broaden our understanding of the mechanism of the generation of ROS in skin layers and the formation/contribution of various excited species can be used as tools to determine the physiological state of the organism.

• Klíčová slova
• malondialdehyde, oxidative radical reaction, porcine skin, protein carbonyls, protein modification, reactive oxygen species, two-dimensional imaging, ultra-weak photon emission,
• MeSH
• kůže * metabolismus   MeSH
• oxidace-redukce MeSH
• oxidační stres MeSH
• peroxid vodíku * metabolismus   MeSH
• proteiny metabolismus   MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• peroxid vodíku * MeSH
• proteiny MeSH
• reaktivní formy kyslíku MeSH

Out of millions of ejaculated sperm, a few reach the fertilization site in mammals. Flagellar Ca2+ signaling nanodomains, organized by multi-subunit CatSper calcium channel complexes, are pivotal for sperm migration in the female tract, implicating CatSper-dependent mechanisms in sperm selection. Here using biochemical and pharmacological studies, we demonstrate that CatSper1 is an O-linked glycosylated protein, undergoing capacitation-induced processing dependent on Ca2+ and phosphorylation cascades. CatSper1 processing correlates with protein tyrosine phosphorylation (pY) development in sperm cells capacitated in vitro and in vivo. Using 3D in situ molecular imaging and ANN-based automatic detection of sperm distributed along the cleared female tract, we demonstrate that spermatozoa past the utero-tubal junction possess the intact CatSper1 signals. Together, we reveal that fertilizing mouse spermatozoa in situ are characterized by intact CatSper channel, lack of pY, and reacted acrosomes. These findings provide molecular insight into sperm selection for successful fertilization in the female reproductive tract.

When mammals mate, males ejaculate millions of sperm cells into the females’ reproductive tract. But as the sperm travel up the tract, only a handful of the ‘fittest’ sperm will actually manage to reach the egg. This process of elimination prevents the egg from being fertilized by multiple sperm cells and stops the eggs from being fertilized outside of the womb. A lot of what is known about fertilization in mammals has come from studying how sperm and eggs cells interact in a Petri dish. However, this approach cannot explain how sperm are selected and removed as they journey towards the egg. Previous work suggests that a calcium channel, which sits in the membrane surrounding the sperm tail, may provide some answers. The core of this channel, known as CatSper, is made up of four proteins arranged into a unique pattern similar to racing stripes. Without this specific arrangement, sperm cells cannot move forward and fertilize the egg in time. To investigate the role of this protein in more depth, Ded et al. established a new way to image the minute structures of sperm cells, such as CatSper, in the reproductive tract of female mice. Experiments in a Petri dish revealed that sperm cells that have been primed to fertilize the egg are a diverse population: in some cells one of the proteins that make up the calcium channel, known as CatSper1, is cleaved, while in other cells this protein remains intact. Visualizing this protein in the female reproductive tract showed that sperm cells close to the site of fertilization contain non-cleaved CatSper1. Whereas sperm cells further away from the egg – and thus closer to the uterus – are more likely to contain broken down CatSper1. Taken together, these findings suggest that the state of the CatSper1 protein may be used to select sperm that are most likely to reach and fertilize the egg. Future studies should address what happens to the calcium channel once the CatSper1 protein is cleaved, and how this channel controls the movements and lifespan of sperm. This could help identify new targets for contraception and improve current strategies for assisted reproduction.

• Klíčová slova
• CatSper, capacitation, cell biology, fertilization, in situ imaging, molecular biophysics, molecular heterogeneity, mouse, sperm, structural biology,
• MeSH
• akrozomální reakce MeSH
• glykosylace MeSH
• molekulární zobrazování metody   MeSH
• myši MeSH
• regulace genové exprese MeSH
• spermie fyziologie   MeSH
• vápníková signalizace MeSH
• vápníkové kanály genetika   metabolismus   MeSH
• ženské pohlavní orgány diagnostické zobrazování   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• Catsper1 protein, mouse MeSH Prohlížeč
• vápníkové kanály MeSH

• MeSH
• azaguanin farmakologie   MeSH
• biofyzika MeSH
• biofyzikální jevy MeSH
• buněčné klony MeSH
• buněčné linie * MeSH
• časové faktory MeSH
• chromozomální aberace MeSH
• genetický kód MeSH
• glukosyltransferasy metabolismus   MeSH
• kultivační techniky MeSH
• kultivované buňky MeSH
• L buňky (buněčná linie) * MeSH
• metody MeSH
• molekulární biologie MeSH
• myši MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• azaguanin MeSH
• glukosyltransferasy MeSH