Molecular dynamics, density functional, ADMET predictions, virtual screening, and molecular interaction field studies for identification and evaluation of novel potential CDK2 inhibitors in cancer therapy

In this work, we have used molecular dynamics, density functional theory, virtual screening, ADMET predictions, and molecular interaction field studies to design and propose eight novel potential inhibitors of CDK2. The eight molecules proposed showed interesting structural characteristics that are required for inhibiting the CDK2 activity and show potential as drug candidates for the treatment of cancer. The parameters related to the Rule of Five were calculated, and only one of the molecules violated more than one parameter. One of the proposals and one of the drug-like compounds selected by virtual screening indicated to be promising candidates for CDK2-based cancer therapy.

Structural complexes of human adenine phosphoribosyltransferase reveal novel features of the APRT catalytic mechanism

Adenine phosphoribosyltransferase (APRT) is an important enzyme component of the purine recycling pathway. Parasitic protozoa of the order Kinetoplastida are unable to synthesize purines de novo and use the salvage pathway for the synthesis of purine bases rendering this biosynthetic pathway an attractive target for antiparasitic drug design. The recombinant human adenine phosphoribosyltransferase (hAPRT) structure was resolved in the presence of AMP in the active site to 1.76 angstrom resolution and with the substrates PRPP and adenine simultaneously bound to the catalytic site to 1.83 angstrom resolution. An additional structure was solved containing one subunit of the dimer in the apo-form to 2.10 angstrom resolution. Comparisons of these three hAPRT structures with other `type I` PRTases revealed several important features of this class of enzymes. Our data indicate that the flexible loop structure adopts an open conformation before and after binding of both substrates adenine and PRPR Comparative analyses presented here provide structural evidence to propose the role of Glu 104 as the residue that abstracts the proton of adenine N9 atom before its nucleophilic attack on the PRPP anomeric carbon. This work leads to new insights to the understanding of the APRT catalytic mechanism.

Use of virtual screening, flexible docking, and molecular interaction fields to design novel HMG-CoA reductase inhibitors for the treatment of hypercholesterolemia

Dietary changes associated with drug therapy can reduce high serum cholesterol levels and dramatically decrease the risk of coronary artery disease, stroke, and overall mortality. Statins are hypolipemic drugs that are effective in the reduction of cholesterol serum levels, attenuating cholesterol synthesis in liver by competitive inhibition regarding the substrate or molecular target HMG-CoA reductase. We have herewith used computer-aided molecular design tools, i.e., flexible docking, virtual screening in large data bases, molecular interaction fields to propose novel potential HMG-CoA reductase inhibitors that are promising for the treatment of hypercholesterolemia.

Molecular dynamics, flexible docking, virtual screening, ADMET predictions, and molecular interaction field studies to design novel potential MAO-B inhibitors

Monoamine oxidase is a flavoenzyme bound to the mitochondrial outer membranes of the cells, which is responsible for the oxidative deamination of neurotransmitter and dietary amines. It has two distinct isozymic forms, designated MAO-A and MAO-B, each displaying different substrate and inhibitor specificities. They are the well-known targets for antidepressant, Parkinson`s disease, and neuroprotective drugs. Elucidation of the x-ray crystallographic structure of MAO-B has opened the way for the molecular modeling studies. In this work we have used molecular modeling, density functional theory with correlation, virtual screening, flexible docking, molecular dynamics, ADMET predictions, and molecular interaction field studies in order to design new molecules with potential higher selectivity and enzymatic inhibitory activity over MAO-B.

Bixin and lycopene modulation of free radical generation induced by cisplatin-DNA interaction

This work evaluated the Modulation of reactive oxygen species (ROS) produced by the cisplatin-human DNA interaction in a cell-free experimental model by the carotenoids bixin and lycopene extracted from, natural dietary Sources and purified through luminol- and Cypridina luciferin methoxy-analogue (MCLA)- enhanced chemiluminescence assays. The results showed that the ROS generation by DNA-cisplatin interaction was inhibited by both lycopene and bixin in a concentration-dependent manner. At a concentration of 100 mu M, lycopene and bixin inhibited Superoxide anion (O center dot(2)) generation at 90% and 82%, respectively, and the total ROS generation at 44% and 42%, respectively. The formation of significant amounts of isomers or degradation products of both carotenoids was not observed after ROS scavenging, as evaluated by high-performance liquid chromatography. Taken together, these results Suggest that carotenoids can be helpful to Modulate the oxidative stress found in cancer therapy with cisplatin. (c) 2008 Elsevier Ltd. All rights reserved.

Biological features of a puzzling symbiotic association between the hermit crab Dardanus insignis and the porcellanid crab Porcellana Sayana (Crustacea)

The symbiotic lifestyle is widespread among porcellanid crabs, which maintain ecological and co-evolutionary associations with annelid polychaetes, poriferans, cnidarians, echinoderms, gastropod. mollusks, and other crustaceans such as shrimps and hermit crabs, among others. We investigated the ecological association between the hermit crab Dardanus insignis and the porcellanid Porcellana sayana, in southeastern Brazil. Porcellanid crabs, hermit crabs, and available shells were collected monthly from July 2001 to June 2003, with a shrimp boat equipped with two double-rig trawl nets. The majority of P. sayana specimens were collected in shells occupied by D. insignis (96.6%); a few were found in empty shells (3.4%). The catch of both symbionts and hosts increased with increasing depth, with the highest occurrence at 35 m. The F. sayana crabs of various sizes could be found solitary or forming aggregations of up to 14 individuals per host, showing no sex or size segregation. In spite of the high diversity of shell species occupied by the hermit crabs and also available in the field, only a few of them were also utilized by P. sayana. The majority (93%) of shells utilized by P. sayana also hosted other symbiont species, constituting the basis of extensive symbiotic complexes. Thus, the ecological relationship between D. insignis and P. sayana may be classified as a non-obligate and non-specific symbiosis that may also involve other facultative organisms such as sea anemones. (C) 2008 Elsevier B.V. All rights reserved.

Real-time monitoring and kinetic parameter estimation of the affinity interaction of jArtinM and rArtinM with peroxidase glycoprotein by the electrogravimetric technique

ArtinM is a D-mannose binding lectin that has been arousing increasing interest because of its biomedical properties, especially those involving the stimulation of Th1 immune response, which confers protection against intracellular pathogens The potential pharmaceutical applications of ArtinM have motivated the production of its recombinant form (rArtinM) so that it is important to compare the sugar-binding properties of jArtinM and rArtinM in order to take better advantage of the potential applications of the recombinant lectin. In this work, a biosensor framework based on a Quartz Crystal Microbalance was established with the purpose of making a comparative study of the activity of native and recombinant ArtinM protein The QCM transducer was strategically functionalized to use a simple model of protein binding kinetics. This approach allowed for the determination of the binding/dissociation kinetics rate and affinity equilibrium constant of both forms of ArtinM with horseradish peroxidase glycoprotein (HRP), a N-glycosylated protein that contains the trimannoside Man alpha 1-3[Man alpha 1-6]Man, which is a known ligand for jArtinM (Jeyaprakash et al, 2004). Monitoring of the real-time binding of rArtinM shows that it was able to bind HRP, leading to an analytical curve similar to that of jArtinM, with statistically equivalent kinetic rates and affinity equilibrium constants for both forms of ArtinM The lower reactivity of rArtinM with HRP than jArtinM was considered to be due to a difference in the number of Carbohydrate Recognition Domains (CRDs) per molecule of each lectin form rather than to a difference in the energy of binding per CRD of each lectin form. (C) 2010 Elsevier B V. All rights reserved

Maternal investment in egg production: environmental and population-specific effects on offspring performance in the snapping shrimp Alpheus nuttingi (Schmitt, 1924) (Decapoda, Alpheidae)

Among marine invertebrates, the overall biomass invested in egg production varies widely within populations, which can result from the interaction of endogenous and exogenous factors. Species that have constant reproduction throughout the year can be good models to study the influence of environmental factors on reproductive processes. We conducted a seasonal comparison of egg production in the intertidal snapping shrimp Alpheus nuttingi, which shows a continuous reproductive pattern, to examine the hypothesis that differences in egg production are driven by environmental conditions and population features. This population showed an uncommon strategy, characterized by females that produce eggs of varying sizes within their clutches, with reduced egg volume when the number of eggs is higher (Spring-Summer). In these seasons, higher temperatures and greater food availability may allow the production of more eggs compared to the Autumn-Winter seasons. Compared to other alpheid shrimps, this population produces small eggs, but in larger numbers. Despite the higher fecundity, the reproductive output is relatively low, this production being supported by the large size of females from the southern Atlantic region. Our findings showed that the egg production of A. nuttingi was greatly influenced by environmental factors. Therefore, this shrimp, and probably other decapods that possess continuous reproduction, adopt different reproductive strategies during the year. (C) Koninklijke Brill NV, Leiden, 2010.

Fight versus flight: the interaction of temperature and body size determines antipredator behaviour in tegu lizards

Ectotherm antipredator behaviour might be strongly affected both by body temperature and size: when environmental temperatures do not favour maximal locomotor performance, large individuals may confront predators, whereas small animals may flee, simply because they have no other option. However, integration of body size and temperature effects is rarely approached in the study of antipredator behaviour in vertebrate ectotherms. In the present study we investigated whether temperature affects antipredator responses of tegu lizards, Tupinambis merianae, with distinct body sizes, testing the hypothesis that small tegus (juveniles) run away from predators regardless of the environmental temperature, because defensive aggression may not be an effective predator deterrent, whereas adults, which are larger, use aggressive defence at low temperatures, when running performance might be suboptimal. We recorded responses of juvenile (small) and adult (large) tegu lizards to a simulated predatory attack at five environmental temperatures in the laboratory. Most differences between the two size classes were observed at low temperatures: large tegus were more aggressive overall than were small tegus at all temperatures tested, but at lower temperatures, the small lizards often used escape responses whereas the large ones either adopted a defensive posture or remained inactive. These results provide strong evidence that body size and temperature affect the antipredator responses of vertebrate ectotherms. We discuss the complex and intricate network of evolutionary and ecological parameters that are likely to be involved in the evolution of such interactions. (C) 2009 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.

Estimating population features of the anomuran crab Petrolisthes armatus (Porcellanidae) in a remaining and impacted mangrove area of the western Atlantic

This study evaluated biological aspects of Petrolisthes armatus inhabiting a remaining but disturbed mangrove area of Brazil. Samples were taken from March 2005 to July 2006, during low tide in the rocky-shore region. The size frequency distribution for all the individuals collected was bimodal. The sex ratio obtained was not different from 1: 1, and the population was classified as standard. A higher reproductive ratio and earlier ovigerous conditions were found in relation to another southern population; these can be hypothesized as adaptations to life in a stressed environment. Even though the population living in Ara has been subject to an environment frequently disturbed by human-produced pollutants, our results show no sign of negative effects on reproductive stages, recruits or members of the population in general. The population profiles of P. armatus show some peculiarities when compared to other populations inhabiting non disturbed environments.

Reproductive strategy of the snapping shrimp Alpheus armillatus H. Milne-Edwards, 1837 in the South Atlantic: fecundity, egg features, and reproductive output

The family Alpheidae, composed by shrimps of relatively small size, popularly known as snapping shrimps, is the one of the most diverse decapod groups. These shrimps are found worldwide and Occur in tropical and subtropical waters, from the intertidal zone to great depths. We investigated reproductive aspects of Alpheus armillatus, in order to gather information on egg production, aiming to enhance knowledge of its reproductive strategies in a population in an intertidal area of the South Atlantic. Ovigerous females were collected under rocks, in May and July 2006 (dry season) and in November 2006 and March 2007 (rainy season). Egg production and reproductive output were analyzed and compared seasonally and during the period of embryonic development. Females measured on average 11.28 mm CL with a mean of 763 eggs and 0.11 mm(3) egg volume. The egg volume of this population was smaller than previous estimates for other species of snapping shrimps, but the mean egg number was higher. The volume of eggs doubled during the incubation period, but despite this increase, no significant loss of eggs was observed. Alpheus armillatus invests oil average about 12% of body weight in reproduction. The proportional investment in egg production IS Significantly higher in the rainy season when compared with the dry season (17.9% vs 4.8%), correlated with higher temperatures and increased food availability at this time. Our results corroborated the hypothesis of a pattern of egg production influenced by environmental conditions and intraspecific variability among the family Alpheidae, as a function of the biogeographic region.

Dermaseptin 01 as antimicrobial peptide with rich biotechnological potential: study of peptide interaction with membranes containing Leishmania amazonensis lipid-rich extract and membrane models

This article addresses the interactions of the synthetic antimicrobial peptide dermaseptin 01 (GLWSTIKQKGKEAAIAAA-KAAGQAALGAL-NH(2), DS 01) with phospholipid (PL) monolayers comprising (i) a lipid-rich extract of Leishmania amazonensis (LRE-La), (ii) zwitterionic PL (dipalmitoylphosphatidylcholine, DPPC), and (iii) negatively charged PL (dipalmitoylphosphatidylglycerol, DPPG). The degree of interaction of DS 01 with the different biomembrane models was quantified from equilibrium and dynamic liquid-air interface parameters. At low peptide concentrations, interactions between DS 01 and zwitterionic PL, as well as with the LRE-La monolayers were very weak, whereas with negatively charged PLs the interactions were stronger. For peptide concentrations above 1 mu g/ml, a considerable expansion of negatively charged monolayers occurred. In the case of DPPC, it was possible to return to the original lipid area in the condensed phase, suggesting that the peptide was expelled from the monolayer. However, in the case of DPPG, the average area per lipid molecule in the presence of DS 01 was higher than pure PLs even at high surface pressures, suggesting that at least part of DS 01 remained incorporated in the monolayer. For the LRE-La monolayers, DS 01 also remained in the monolayer. This is the first report on the antiparasitic activity of AMPs using Langmuir monolayers of a natural lipid extract from L. amazonensis. Copyright (C) 2011 European Peptide Society and John Wiley & Sons, Ltd.

Porphyrin-phospholipid interaction and ring metallation depending on the phospholipid polar head type

The interaction between a hydrophobically modified 5,10,15,20-tetrakis(4-N-tetradecyl-pyridyl) porphyrin and three phospholipids: two negatively charged, DMPA (the sodium salt of dimyristoyl-sn-glycero-phosphatidyl acid) and DMPG (the sodium salt of 1,2-dimyristoyl-sn-glycero-3-[phospho-rac-(1-glycerol)]) and a zwitterionic DMPC (dimyristoyl-sn-glycero-phosphatidylcholine), were studied by means of surface pressure isotherms and spectroscopic methods. The interaction results in partial or total metallation of the porphyrin with zinc ions in the presence of negatively charged phospholipids, as attested by UV-vis and luminescence spectroscopy of the transferred films. In the presence of the zwitterionic phospholipid no insertion of zinc ion in the porphyrin ring is detected. These results are relevant for the understanding of photosensitizer-lipid-carrier binding for use in photodynamic therapy. (C) 2010 Elsevier Inc. All rights reserved.

Interaction of 10-(octyloxy) decyl-2-(trimethylammonium) ethyl phosphate with mimetic membranes and cytotoxic effect on leukemic cells

10-(Octyloxy) decyl-2-(trimethylammonium) ethyl phosphate (ODPC) is an alkylphospholipid that can interact with cell membranes because of its amphiphilic character. We describe here the interaction of ODPC with liposomes and its toxicity to leukemic cells with an ED-50 of 5.4, 5.6 and 2.9 pM for 72 h of treatment for inhibition of proliferation of NB4, U937 and K562 cell lines, respectively, and lack of toxicity to normal hematopoietic progenitor cells at concentrations up to 25 pM. The ED-50 for the non-malignant HEK-293 and primary human umbilical vein endothelial cells (HUVEC) was 63.4 and 60.7 mu M, respectively. The critical micellar concentration (CMC) of ODPC was 200 mu M. Dynamic light scattering indicated that dipalmitoylphosphatidylcholine (DPPC) liposome size was affected only above the CMC of ODPC. Differential calorimetric scanning (DCS) of liposomes indicated a critical transition temperature (T(c)) of 41.5 degrees C and an enthalpy (Delta H) variation of 7.3 kcal mol(-1). The presence of 25 mu M ODPC decreased T(c) and Delta H to 393 degrees C and 4.7 kcal mol(-1), respectively. ODPC at 250 mu M destabilized the liposomes (36.3 degrees C. 0.46 kcal mol(-1)). Kinetics of 5(6)-carboxyfluorescein (CF) leakage from different liposome systems indicated that the rate and extent of CF release depended on liposome composition and ODPC concentration and that above the CMC it was instantaneous. Overall, the data indicate that ODPC acts on in vitro membrane systems and leukemia cell lines at concentrations below its CMC, suggesting that it does not act as a detergent and that this effect is dependent on membrane composition. (C) 2010 Elsevier B.V. All rights reserved.

The interaction of bothropstoxin-I (Lys49-PLA(2)) with liposome membranes

Bothropstoxin-I (BthTx-I) is a Lys49-PLA(2) from the venom of the snake Bothrops jararacussu, which permeabilizes biological and artificial membranes by a mechanism independent of lipid hydrolysis. This mechanism has been investigated by studying the interaction of nine single tryptophan BthTx-I mutants with negatively charged phospholipid membranes. Changes in the solvent exposure of the tryptophan in each mutant were evaluated comparing the rate of chemical modification (k(mod)) by bromosuccinamide with the maximum intrinsic tryptophan fluorescence emission wavelength (lambda(max)) in buffer and in the presence of 10% DMPA/90% DPPC liposomes. No changes in lambda(max). were observed, whereas k(mod) values for tryptophans at positions 7, 10, 31 and 125 were significantly reduced in the presence of lipids, suggesting that bound phospholipid decreases solvent accessibility at these positions. Since the half-lives of the fluorescence and chemical modification effects differ by at least six orders of magnitude, these results suggest that the bound phospholipid may interact with multiple locations on the protein surface over micro- to millisecond timescales. (C) 2009 Elsevier Ltd. All rights reserved.