DNA-Interactive Properties of Crotamine, a Cell-Penetrating Polypeptide and a Potential Drug Carrier

Crotamine, a 42-residue polypeptide derived from the venom of the South American rattlesnake Crotalus durissus terrificus, has been shown to be a cell-penetrating protein that targets chromosomes, carries plasmid DNA into cells, and shows specificity for actively proliferating cells. Given this potential role as a nucleic acid-delivery vector, we have studied in detail the binding of crotamine to single- and double-stranded DNAs of different lengths and base compositions over a range of ionic conditions. Agarose gel electrophoresis and ultraviolet spectrophotometry analysis indicate that complexes of crotamine with long-chain DNAs readily aggregate and precipitate at low ionic strength. This aggregation, which may be important for cellular uptake of DNA, becomes less likely with shorter chain length. 25-mer oligonucleotides do not show any evidence of such aggregation, permitting the determination of affinities and size via fluorescence quenching experiments. The polypeptide binds non-cooperatively to DNA, covering about 5 nucleotide residues when it binds to single (ss) or (ds) double stranded molecules. The affinities of the protein for ss-vs. ds-DNA are comparable, and inversely proportional to salt levels. Analysis of the dependence of affinity on [NaCl] indicates that there are a maximum of,3 ionic interactions between the protein and DNA, with some of the binding affinity attributable to non-ionic interactions. Inspection of the three-dimensional structure of the protein suggests that residues 31 to 35, Arg-Trp-Arg-Trp-Lys, could serve as a potential DNA-binding site. A hexapeptide containing this sequence displayed a lower DNA binding affinity and salt dependence as compared to the full-length protein, likely indicative of a more suitable 3D structure and the presence of accessory binding sites in the native crotamine. Taken together, the data presented here describing crotamine-DNA interactions may lend support to the design of more effective nucleic acid drug delivery vehicles which take advantage of crotamine as a carrier with specificity for actively proliferating cells. Citation: Chen P-C, Hayashi MAF, Oliveira EB, Karpel RL (2012) DNA-Interactive Properties of Crotamine, a Cell-Penetrating Polypeptide and a Potential Drug Carrier. PLoS ONE 7(11): e48913. doi:10.1371/journal.pone.0048913

Revision and cladistic analysis of Neotropical genus Psilochlorops Duda (Diptera: Chloropidae)

Five new species are described herein for the Neotropical genus Psilochlorops, up to now known only from the type-species, P. clavitibia Duda-P. brunneus sp. n., P. elongatum sp. n., P. flavisoma sp. n., P. nigrifemur sp. n. and P. paganelliae sp. n. All new species are described in detail and illustrated. A key to the species of the genus is provided. The diagnosis of Psilochlorops, after the addition of these species, is emended. A cladistic analysis of the genus indicates that Psilochlorops is monophyletic and show the affinities between the species.

A New Species of Cascudinho of the Genus Hisonotus (Siluriformes: Loricariidae: Hypoptopomatinae) from the Upper Rio Tapajos Basin, Brazil

Hisonotus bockmanni, new species, is described based on specimens collected in a sandbank in the Rio Cururu, a tributary to the Rio Teles Pires, one of the rivers forming the Rio Tapajos in the Amazon Basin. The new taxon is distinguished from its congeners by a unique color pattern, whose most striking features are: two elliptical white spots, anterior to nostrils; predorsal region darkly pigmented with five unpigmented spots arranged as anteriorly pointed chevron; and a rostrocaudally elongate cross along most of the caudal peduncle. The placement of the new species in Hisonotus as well as its possible affinities within that genus are discussed in light of the current knowledge of the phylogenetic relationships among the Hypoptopomatinae.

Calcium Binding to Leptospira Outer Membrane Antigen LipL32 Is Not Necessary for Its Interaction with Plasma Fibronectin, Collagen Type IV, and Plasminogen

LipL32 is the most abundant outer membrane protein from pathogenic Leptospira and has been shown to bind extracellular matrix (ECM) proteins as well as Ca2+. Recent crystal structures have been obtained for the protein in the apo-and Ca2+-bound forms. In this work, we produced three LipL32 mutants (D163-168A, Q67A, and S247A) and evaluated their ability to interact with Ca2+ and with ECM glycoproteins and human plasminogen. The D163-168A mutant modifies aspartate residues involved in Ca2+ binding, whereas the other two modify residues in a cavity on the other side of the protein structure. Loss of calcium binding in the D163-D168A mutant was confirmed using intrinsic tryptophan fluorescence, circular dichroism, and thermal denaturation whereas the Q67A and S247A mutants presented the same Ca2+ affinity as the wild-type protein. We then evaluated if Ca2+ binding to LipL32 would be crucial for its interaction with collagen type IV and plasma proteins fibronectin and plasminogen. Surprisingly, the wild-type protein and all three mutants, including the D163-168A variant, bound to these ECM proteins with very similar affinities, both in the presence and absence of Ca2+ ions. In conclusion, calcium binding to LipL32 may be important to stabilize the protein, but is not necessary to mediate interaction with host extracellular matrix proteins.

Dihydrocinnamic acid derivatives from Hortia species and their chemotaxonomic value in the Rutaceae

Phytochemical studies of Hortia brasiliana and Hortia oreadica (Rutaceae) have led to the identification of three novel dihydrocinnamic acids: 5-methoxy-2,2-dimethyl-2H-1-benzopyran-8-propanoic acid, 5,6-dimethoxy-2,2-dimethyl-2H-1-benzopyran-8-propanoic acid and erythro-2-hydroxy-4-methoxy-3-(1,2,3-trihydroxy-3-methylbutyl) benzenepropanoic acid from H. brasiliana and the second compound and six known dihydrocinnamic acids from H. oreadica. Engler included Hortia as the single Neotropical genus in the Toddalioideae subtribe Toddaliinae. However, the range of dihydrocinnamic acid derivatives found in H. brasiliana and H. oreadica show that they contain similar compounds to other species of Hortia and clearly point to their phytochemical affinities with other Rutoideae species. (C) 2012 Elsevier Ltd. All rights reserved.

Fishes of Paranagua Estuarine Complex, South West Atlantic

The objective of this work was to present an updated checklist of the currently known fishes in the Paranagua Estuarine Complex (PEC) and provides comments on conservation status for the treated species. We used a large dataset derived from a pool of studies which have been conducted within there along the last 30 years. Each study was based on monthly samplings and conducted in several estuarine habitat; thus, the pool covers practically all estuarine habitats and takes into account the seasonal cycle in the system. The PEC ichthyofauna represents a mixture between that fauna typical from the tropical Brazilian coast and that with affinities of temperate Argentinean and Uruguayan zones. The PEC harbors a rich fish fauna of 213 species, inserted in the families that are common along the Brazilian coast. Only a minor part (8%) of the PEC fish fauna was evaluated as regards the conservation status, mostly because of the lack of basic biological and ecological information for most species. Despite part of the among-estuaries differences are due to different and incomplete sampling efforts, the richness in the PEC is surprisingly higher than other systems in Brazil and around world, which emphasize the importance of the region for global biodiversity conservation.

LAGOA SANTA REVISITED: AN OVERVIEW OF THE CHRONOLOGY, SUBSISTENCE, AND MATERIAL CULTURE OF PALEOINDIAN SITES IN EASTERN CENTRAL BRAZIL

Lagoa Santa, a karstic area in eastern Central Brazil, has been subject to research on human paleontology and archaeology for 175 years. Almost 300 Paleoindian human skeletons have been found since Danish naturalist Peter Lund's pioneering work. Even so, some critical issues such as the role of rockshelters in settlement systems, and the possible paleoclimatic implications of the peopling of the region have yet to be addressed. We present some results obtained from recent excavations at four rockshelters and two open-air sites, new dates for human Paleoindian skeletons, and a model to explain the cultural patterns observed so far. It is also argued that the Paleoindian subsistence system at Lagoa Santa was similar to other locations in South America: generalized small-game hunting complemented by fruits, seed, and root gathering.

Novel organization of the mitochondrial genome in the deep-sea coral, Madrepora oculata (Hexacorallia, Scleractinia, Oculinidae) and its taxonomic implications

Madrepora is one of the most ecologically important genera of reef-building scleractinians in the deep sea, occurring from tropical to high-latitude regions. Despite this, the taxonomic affinities and relationships within the genus Madrepora remain unclear. To clarify these issues, we sequenced the mitochondrial (mt) genome of the most widespread Madrepora species, M. oculata, and compared this with data for other scleractinians. The architecture of the M. oculara mt genome was very similar to that of other scleractinians, except for a novel gene rearrangement affecting only cox2 and cox3. This pattern of gene organization was common to four geographically distinct M. oculata individuals as well as the congeneric species M. minutiseptum, but was not shared by other genera that are closely related on the basis of cox1 sequence analysis nor other oculinids, suggesting that it might be unique to Madrepora. (C) 2012 Elsevier Inc. All rights reserved.

Characterization of suramin binding sites on the human group IIA secreted phospholipase A(2) by site-directed mutagenesis and molecular dynamics simulation

Suramin is a polysulphonated naphthylurea with inhibitory activity against the human secreted group IIA phospholipase A(2) (hsPLA2GIIA), and we have investigated suramin binding to recombinant hsPLA2GIIA using site-directed mutagenesis and molecular dynamics (MD) simulations. The changes in suramin binding affinity of 13 cationic residue mutants of the hsPLA2GIIA was strongly correlated with alterations in the inhibition of membrane damaging activity of the protein. Suramin binding to hsPLA2GIIA was also studied by MD simulations, which demonstrated that altered intermolecular potential energy of the suramin/mutant complexes was a reliable indicator of affinity change. Although residues in the C-terminal region play a major role in the stabilization of the hsPLA2GIIA/suramin complex, attractive and repulsive hydrophobic and electrostatic interactions with residues throughout the protein together with the adoption of a bent suramin conformation, all contribute to the stability of the complex. Analysis of the h5PLA2GIIA/suramin interactions allows the prediction of the properties of suramin analogues with improved binding and higher affinities which may be candidates for novel phospholipase A(2) inhibitors. (C) 2012 Elsevier Inc. All rights reserved.

Placentation in dolphins from the Amazon River Basin: the Boto, Inia geoffrensis, and the Tucuxi, Sotalia fluviatilis

A recent reassessment of the phylogenetic affinities of cetaceans makes it timely to compare their placentation with that of the artiodactyls. We studied the placentae of two sympatric species of dolphin from the Amazon River Basin, representing two distinct families. The umbilical cord branched to supply a bilobed allantoic sac. Small blood vessels and smooth muscle bundles were found within the stroma of the cord. Foci of squamous metaplasia occurred in the allanto-amnion and allantochorion. The interhemal membrane of the placenta was of the epitheliochorial type. Two different types of trophoblastic epithelium were seen. Most was of the simple columnar type and indented by fetal capillaries. However, there were also areolar regions with tall columnar trophoblast and these were more sparsely supplied with capillaries. The endometrium was well vascularised and richly supplied with actively secreting glands. These findings are consistent with the current view that Cetacea are nested within Artiodactyla as sister group to the hippopotamids.

Phylogenetic relationships of the genus Sibynophis (Serpentes: Colubroidea)

We present the results of the first molecular analysis of the phylogenetic affinities of the Asian colubroid genus Sibynophis. We recovered a sister-group relationship between Sibynophis and the New World Scaphiodontophis. Although Liophidium sometimes is associated with these genera, the relationship is distant. Morphological characters that Liophidium shares with Sibynophis and Scaphiodontophis are resolved as homoplasies that probably reflect the similarities of their specialized feeding habits. The traditional subfamily Sibynophiinae is elevated to the family-level, and Scaphiodontophiinae is placed in its synonymy.

Fishes of Paranaguá Estuarine Complex, South West Atlantic

The objective of this work was to present an updated checklist of the currently known fishes in the Paranaguá Estuarine Complex (PEC) and provides comments on conservation status for the treated species. We used a large dataset derived from a pool of studies which have been conducted within there along the last 30 years. Each study was based on monthly samplings and conducted in several estuarine habitat; thus, the pool covers practically all estuarine habitats and takes into account the seasonal cycle in the system. The PEC ichthyofauna represents a mixture between that fauna typical from the tropical Brazilian coast and that with affinities of temperate Argentinean and Uruguayan zones. The PEC harbors a rich fish fauna of 213 species, inserted in the families that are common along the Brazilian coast. Only a minor part (8%) of the PEC fish fauna was evaluated as regards the conservation status, mostly because of the lack of basic biological and ecological information for most species. Despite part of the among-estuaries differences are due to different and incomplete sampling efforts, the richness in the PEC is surprisingly higher than other systems in Brazil and around world, which emphasize the importance of the region for global biodiversity conservation.

Louis-Bernard Guyton de Morveau e a revolução química das Luzes

O objetivo deste artigo é investigar a concepção enciclopédica de revolução científica posta em prática pelo químico francês L.-B. Guyton de Morveau (1737-1816). Deslocando a análise do conhecimento químico das Luzes do programa traçado por Lavoisier (1743-1794), sugerimos uma concepção revolucionária republicana, proclamada como resultado do esforço de uma coletividade. Daremos destaque a três abordagens revolucionárias de Guyton de Morveau no âmbito da química. A primeira foi sua atuação no ensino dessa ciência, cuja pedagogia e métodos de ensino foram fundamentais para sua imersão social. Além disso, entre 1770 e 1790, Guyton de Morveau teve desempenhos decisivos no seio da empresa enciclopédica e no seio da escola química francesa.

Deltocyathiidae, an early-diverging family of Robust corals (Anthozoa, Scleractinia)

Over the last decade, molecular phylogenetics has called into question some fundamental aspects of coral systematics. Within the Scleractinia, most families composed exclusively by zooxanthellate species are polyphyletic on the basis of molecular data, and the second most speciose coral family, the Caryophylliidae (most members of which are azooxanthellate), is an unnatural grouping. As part of the process of resolving taxonomic affinities of caryophylliids', here a new Robust' scleractinian family (Deltocyathiidae fam. n.) is proposed on the basis of combined molecular (CO1 and 28S rDNA) and morphological data, accommodating the early-diverging clade of traditional caryophylliids (represented today by the genus Deltocyathus). Whereas this family captures the full morphological diversity of the genus Deltocyathus, one species, Deltocyathus magnificus, is an outlier in terms of molecular data, and groups with the Complex coral family Turbinoliidae. Ultrastructural data, however, place D.magnificus within Deltocyathiidae fam. nov. Unfortunately, limited ultrastructural data are as yet available for turbinoliids, but D.magnificus may represent the first documented case of morphological convergence at the microstructural level among scleractinian corals. Marcelo V.Kitahara, Centro de Biologia Marinha, Universidade de SAo Paulo, SAo SebastiAo, S.P. 11600-000, Brazil. E-mail:kitahara@usp.br

Synthese, n.c.a. 18F-Markierung und Evaluierung von Antagonisten der Strychnin-unempfindlichen-Glycinbindungsstelle auf Basis der 4-Amino-5,7-Dichlor-1,2,3,4-Tetrahydro-Chinolin-2-Carbonsäure

ZUSAMMENFASSUNGDer glutamaterge N-Methyl-D-aspartat-Rezeptor (NMDA) ist ein wichtiger ionotroper Rezeptor, der die exzitatorische synaptische Transmission im zentralen Nervensystem von Säugetieren vermittelt. Der NMDA-Rezeptor nimmt unter den Glutamatrezeptoren dabei eine Sonderstellung ein, da er mit einer Reihe von neurodegenerativen Erkrankungen wie dem Morbus Parkinson, dem Morbus Huntington, dem Morbus Alzheimer, der Schizophrenie und der Epilepsie in Zusammenhang gebracht wird. Daher besteht ein großes Interesse an der Entwicklung geeigneter 18F-markierter NMDA-Rezeptorliganden zur nicht-invasiven Visualisierung des NMDA-Rezeptorkomplexes mittels der Positronenemissionstomographie.Die 19F-Analoga ADTC1, tADTC1 und tADTC3 - 5 und das nicht-fluorierte 12C-Analogon tADTC2 wurden synthetisiert und ihre in-vitro Affinität und Lipophilie bestimmt. Mit Ausnahme von ADTC1 und tADTC5 die mikromolare Affinitäten besitzen, haben die Liganden in [H-3]MDL-105,519 Rezeptorbindungsassays niedrige nanomolare Affinitäten für die Glycinbindungsstelle. Die Lipophilie der Verbindungen wurde mit drei verschiedenen Verfahren untersucht und ergab logD7,4-Werte von ungefähr 1 für cADTC1 und tADTC1 – 4, während tADTC5 mit einem logD7,4 von –1,15 eine sehr niedrige Lipophilie aufwies. Die Radiosynthesen der 18F-Liganden wurden hinsichtlich der Umsetzung der Markierungsvorläufer mit 2-[F-18]Fluorethyltosylat oder [F-18]Fluorid untersucht und optimiert. Die höchsten radiochemischen Ausbeuten von ungefähr 90% wurden, unter Verwendung von NaOH als Hilfsbase, bei der 18F-Fluorethylierung von t[F-18]ADTC4 und t[F-18]ADTC5 mit 2-[F-18]Fluorethyltosylat erzielt.