VCD to determine absolute configuration of natural product molecules: secolignans from Peperomia blanda

The absolute configuration and solution-state conformers of three peperomin-type secolignans isolated from Peperomia blanda (Piperaceae) are unambiguously determined by using vibrational circular dichroism (VCD) spectroscopy associated with density functional theory (DFT) calculations. Advantages of VCD over the electronic form of CD for the analysis of diastereomers are also discussed. This work extends our growing knowledge about secondary metabolites within the Piperaceae family species while providing a definitive and straightforward method to assess the absolute stereochemistry of secolignans.

Distribution of human immunodeficiency virus type 1 subtypes in the State of Amazonas, Brazil, and subtype C identification

Few studies have reported the molecular epidemiological characterization of HIV-1 in the Northern region of Brazil. The present study reports the molecular and epidemiological characterization of 31 HIV-1 isolates from blood donors from the State of Amazonas who donated blood between April 2006 and March 2007. Serum/plasma samples from all donors were screened for HIV antibodies by ELISA and the results confirmed by Western blot analysis. Genomic DNA was extracted from the buffy coat using the Super Quik-Gene-DNA Isolation kit. Nested PCR was performed on the env, gag, and pol regions of HIV-1 using the Gene Amp PCR System 9700. Sequencing reactions were performed using the inner PCR primers and the DYEnamic (TM) ET Dye Terminator Kit, and phylogenetic analysis was performed using the gag, pol, and env gene sequences. We collected samples from 31 blood donors who tested positive for HIV-1 in confirmatory experiments. The male: female ratio of blood donors was 3.4:1, and the mean age was 32.4 years (range: 19 to 61 years). Phylogenetic analysis showed that subtype B is the most prevalent among Northern Brazilian HIV-1-seropositive blood donors. One HIV-1 subtype C and one circulating recombinant form (CRF_BF) of HIV-1 were identified in the State of Amazonas. This is the first study showing the occurrence of a possible "homogenous" subtype C in this region of Brazil. This finding could contribute to a better characterization of the HIV-1 strains that circulate in the country.

Influence of milk type and addition of passion fruit peel powder on fermentation kinetics, texture profile and bacterial viability in probiotic yoghurts

The effect of the addition of passion fruit peel powder (PFPP) on the fermentation kinetics and texture parameters, post-acidification and bacteria counts of probiotic yoghurts made with two milk types were evaluated during 28 days of storage at 4 degrees C. Milks were fermented by Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus (CY340), and one strain of probiotic bacteria: Lactobacillus acidophilus (L10 and NCFM), Bifidobacterium animalis subsp. lactis (8104 and HN019). The addition of PFPP reduced significantly fermentation time of skim milk co-fermented by the strains L10, NCFM and HN019. At the end of 28-day shelf-life, counts of B. lactis Bl04 were about 1 Log CFU mL(-1) higher in whole yoghurt fermented with PFPP regarding its control but, in general, the addition of PFPP had less influence on counts than the milk type itself. The titratable acidity in yoghurts with PFPP was significantly higher than in their respective controls, and in skim yoghurts higher than in the whole ones. The PFPP increased firmness, consistency (except for the NCFM strain of L acidophilus) and cohesiveness of all skim yoghurts. The results point out the suitability of using passion fruit by-product in the formulation of both skim and whole probiotic yoghurts. (C) 2012 Elsevier Ltd. All rights reserved.

Distribution of human immunodeficiency virus type 1 subtypes in the state of Amazonas, Brazil, and subtype C identification

Few studies have reported the molecular epidemiological characterization of HIV-1 in the Northern region of Brazil. The present study reports the molecular and epidemiological characterization of 31 HIV-1 isolates from blood donors from the State of Amazonas who donated blood between April 2006 and March 2007. Serum/plasma samples from all donors were screened for HIV antibodies by ELISA and the results confirmed by Western blot analysis. Genomic DNA was extracted from the buffy coat using the Super Quik-Gene-DNA Isolation kit. Nested PCR was performed on the env, gag, and pol regions of HIV-1 using the Gene Amp PCR System 9700. Sequencing reactions were performed using the inner PCR primers and the DYEnamic™ ET Dye Terminator Kit, and phylogenetic analysis was performed using the gag, pol, and env gene sequences. We collected samples from 31 blood donors who tested positive for HIV-1 in confirmatory experiments. The male:female ratio of blood donors was 3.4:1, and the mean age was 32.4 years (range: 19 to 61 years). Phylogenetic analysis showed that subtype B is the most prevalent among Northern Brazilian HIV-1-seropositive blood donors. One HIV-1 subtype C and one circulating recombinant form (CRF_BF) of HIV-1 were identified in the State of Amazonas. This is the first study showing the occurrence of a possible "homogenous" subtype C in this region of Brazil. This finding could contribute to a better characterization of the HIV-1 strains that circulate in the country.

Structure of the PilZ-FimXEAL-c-di-GMP complex responsible for the regulation of bacterial type IV pilus biogenesis

Signal transduction pathways mediated by cyclic-bis(3'→5')-dimeric GMP (c-di-GMP) control many important and complex behaviors in bacteria. C-di-GMP is synthesized through the action of GGDEF domains that possess diguanylate cyclase activity and is degraded by EAL or HD-GYP domains with phosphodiesterase activity. There is mounting evidence that some important c-di-GMP-mediated pathways require protein-protein interactions between members of the GGDEF, EAL, HD-GYP and PilZ protein domain families. For example, interactions have been observed between PilZ and the EAL domain from FimX of Xanthomonas citri (Xac). FimX and PilZ are involved in the regulation of type IV pilus biogenesis via interactions of the latter with the hexameric PilB ATPase associated with the bacterial inner membrane. Here, we present the crystal structure of the ternary complex made up of PilZ, the FimX EAL domain (FimXEAL) and c-di-GMP. PilZ interacts principally with the lobe region and the N-terminal linker helix of the FimXEAL. These interactions involve a hydrophobic surface made up of amino acids conserved in a non-canonical family of PilZ domains that lack intrinsic c-di-GMP binding ability and strand complementation that joins β-sheets from both proteins. Interestingly, the c-di-GMP binds to isolated FimXEAL and to the PilZ-FimXEAL complex in a novel conformation encountered in c-di-GMP-protein complexes in which one of the two glycosidic bonds is in a rare syn conformation while the other adopts the more common anti conformation. The structure points to a means by which c-di-GMP and PilZ binding could be coupled to FimX and PilB conformational states

Development of Organocatalytic stereoselective SN1 type reactions

The proposal in my thesis has been the study of Stereoselective α-alkylation through SN1 type reaction. SN1 type reaction involves a stabilized and reactive carbocation intermediate By taking advantages of stability of particular carbocations, the use of carbocations in selective reactions has been important. In this work has been necessary to know the stability and reactivity of carbocations. And the work of Mayr group has helped to rationalize the behaviour and reactivity between the carbocations and nucleophiles by the use of Mayr’s scale of reactivity. The use of alcohols to performed the stable and reactive carbocations have been the key in my thesis. The direct nucleophilic substitution of alcohols has been a crucial scope in the field of organic synthesis, because offer a wide range of intermediates for the synthesis of natural products and pharmaceutics synthesis. In particular the catalytic nucleophilic direct substitution of alcohols represents a novel methodology for the preparation of a variety of derivatives, and water only as the sub-product in the reaction. The stereochemical control of the transformation C-H bond into stereogenic C-C bond adjacent to carbonyl functionalized has been studied for asymmetric catalysis. And the field of organocatalysis has introduced the use of small organic molecule as catalyst for stereoselective transformations. Merging these two concepts Organocatalysis and Mayr’s scale, my thesis has developed a new approach for the α-alkylation of aldehydes and ketones through SN1 type reaction.

The group of invariants of an inner function with finite spectrum

This paper determines the group of continuous invariants corresponding to an inner function circle dot with finitely many singularities on the unit circle T; that is, the continuous mappings g : T -> T such that circle dot o g = circle dot on T. These mappings form a group under composition.

Prime and Semiprime Inner Functions

This paper studies the structure of inner functions under the operation of composition, and in particular the notions or primeness and semiprimeness. Results proved include the density of prime finite Blaschke products in the set of finite Blaschke products, the semiprimeness of finite products of thin Blaschke products and their approximability by prime Blaschke products. An example of a nonsemiprime Blaschke product that is a Frostman Blaschke product is also provided.

Prime and semiprime inner functions

Abstract This paper studies the structure of inner functions under the operation of composition, and in particular the notions or primeness and semiprimeness. Results proved include the density of prime finite Blaschke products in the set of finite Blaschke products, the semiprimeness of finite products of thin Blaschke products and their approximability by prime Blaschke products. An example of a nonsemiprime Blaschke product that is a Frostman Blaschke product is also provided.

Fic domain-catalyzed adenylylation: insight provided by the structural analysis of the type IV secretion system effector BepA

Numerous bacterial pathogens subvert cellular functions of eukaryotic host cells by the injection of effector proteins via dedicated secretion systems. The type IV secretion system (T4SS) effector protein BepA from Bartonella henselae is composed of an N-terminal Fic domain and a C-terminal Bartonella intracellular delivery domain, the latter being responsible for T4SS-mediated translocation into host cells. A proteolysis resistant fragment (residues 10-302) that includes the Fic domain shows autoadenylylation activity and adenylyl transfer onto Hela cell extract proteins as demonstrated by autoradiography on incubation with α-[(32)P]-ATP. Its crystal structure, determined to 2.9-Å resolution by the SeMet-SAD method, exhibits the canonical Fic fold including the HPFxxGNGRxxR signature motif with several elaborations in loop regions and an additional β-rich domain at the C-terminus. On crystal soaking with ATP/Mg(2+), additional electron density indicated the presence of a PP(i) /Mg(2+) moiety, the side product of the adenylylation reaction, in the anion binding nest of the signature motif. On the basis of this information and that of the recent structure of IbpA(Fic2) in complex with the eukaryotic target protein Cdc42, we present a detailed model for the ternary complex of Fic with the two substrates, ATP/Mg(2+) and target tyrosine. The model is consistent with an in-line nucleophilic attack of the deprotonated side-chain hydroxyl group onto the α-phosphorus of the nucleotide to accomplish AMP transfer. Furthermore, a general, sequence-independent mechanism of target positioning through antiparallel β-strand interactions between enzyme and target is suggested.

The IS/OS junction layer in the natural history of type 2 idiopathic macular telangiectasia

To document the progression of a break in the photoreceptor inner segment/outer segment (IS/OS) junction layer and its functional correlates over time in the natural history of type 2 idiopathic macular telangiectasia (type 2 MacTel).

"En face" OCT imaging of the IS/OS junction line in type 2 idiopathic macular telangiectasia

We investigated abnormalities of the photoreceptor inner/outer segment (IS/OS) junction layer viewed "en face" and their functional correlates in type 2 idiopathic macular telangiectasia (type 2 MacTel).

An essential bacterial-type cardiolipin synthase mediates cardiolipin formation in a eukaryote

Cardiolipin is important for bacterial and mitochondrial stability and function. The final step in cardiolipin biosynthesis is catalyzed by cardiolipin synthase and differs mechanistically between prokaryotes and eukaryotes. To study the importance of cardiolipin synthesis for mitochondrial integrity, membrane protein complex formation, and cell proliferation in the human and animal pathogenic protozoan parasite, Trypanosoma brucei, we generated conditional cardiolipin synthase-knockout parasites. We found that cardiolipin formation in T. brucei procyclic forms is catalyzed by a bacterial-type cardiolipin synthase, providing experimental evidence for a prokaryotic-type cardiolipin synthase in a eukaryotic organism. Ablation of enzyme expression resulted in inhibition of de novo cardiolipin synthesis, reduction in cellular cardiolipin levels, alterations in mitochondrial morphology and function, and parasite death in culture. By using immunofluorescence microscopy and blue-native gel electrophoresis, cardiolipin synthase was shown to colocalize with inner mitochondrial membrane proteins and to be part of a large protein complex. During depletion of cardiolipin synthase, the levels of cytochrome oxidase subunit IV and cytochrome c1, reflecting mitochondrial respiratory complexes IV and III, respectively, decreased progressively.

Why is 11beta-hydroxysteroid dehydrogenase type 1 facing the endoplasmic reticulum lumen? Physiological relevance of the membrane topology of 11beta-HSD1

11Beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) is essential for the local activation of glucocorticoid receptors (GR). Unlike unliganded cytoplasmic GR, 11beta-HSD1 is an endoplasmic reticulum (ER)-membrane protein with lumenal orientation. Cortisone might gain direct access to 11beta-HSD1 by free diffusion across membranes, indirectly via intracellular binding proteins or, alternatively, by insertion into membranes. Membranous cortisol, formed by 11beta-HSD1 at the ER-lumenal side, might then activate cytoplasmic GR or bind to ER-lumenal secretory proteins. Compartmentalization of 11beta-HSD1 is important for its regulation by hexose-6-phosphate dehydrogenase (H6PDH), which regenerates cofactor NADPH in the ER lumen and stimulates oxoreductase activity. ER-lumenal orientation of 11beta-HSD1 is also essential for the metabolism of the alternative substrate 7-ketocholesterol (7KC), a major cholesterol oxidation product found in atherosclerotic plaques and taken up from processed cholesterol-rich food. An 11beta-HSD1 mutant adopting cytoplasmic orientation efficiently catalyzed the oxoreduction of cortisone but not 7KC, indicating access to cortisone from both sides of the ER-membrane but to 7KC only from the lumenal side. These aspects may be relevant for understanding the physiological role of 11beta-HSD1 and for developing therapeutic interventions to control glucocorticoid reactivation.

Autosomal-dominant isolated growth hormone deficiency (IGHD type II) with normal GH-1 gene

BACKGROUND: Autosomal-dominant isolated GH deficiency (IGHD) is a rare disorder that is commonly believed to be due to heterozygous mutations in the GH-1 gene (GH-1). These mutations cause the production of a protein that affects the release of the product of the normal allele. Rarely, heterozygous mutations in the gene encoding for HESX-1 gene (HESX-1) may cause autosomal-dominant IGHD, with penetrance that has been shown to be variable in both humans and mice. SUBJECTS AND METHODS: We have sequenced the whole GH-1 in the index cases of 30 families with autosomal-dominant IGHD. In all the families other possible causes of GH deficiency and other pituitary hormones deficits were excluded. We here describe the clinical, biochemical and radiological picture of the families without GH-1 mutations. In these families, we also sequenced the HESX-1. RESULTS: The index cases of the five families with autosomal-dominant IGHD had normal GH-1, including the intronic sequences. They had no HESX-1 mutations. CONCLUSION: This study shows that GH-1 mutations are absent in 5/30 (16.6%) of the families with autosomal-dominant IGHD and raises the possibility that mutations in other gene(s) may be involved in IGHD with this mode of transmission.