Aluminum chloride-mediated kinetic resolution of racemic gamma-substituted-gamma-lactones

Preparation of chiral gamma-substituted-gamma-lactones (1) through kinetic resolution is described. (S)-(-)-1-Phenylethylamine (2) in the presence of anhydrous AlCl(3) shows satisfactory levels of enantioselection in reaction with racemic gamma-substituted-gamma-lactones 1, where (R)-1 remains unreacted, while (S)-1 is enantioselectively converted to the ring-opened amide (S,S)-4. The enantiopurity of (R)-(+)- gamma-substituted gamma-lactones recovered ranges from 62-98% ee.     

Modulation of pigment profiles of Calothrix elenkenii in response to environmental changes

Cyanobacteria are versatile tetrapyrrole synthesizers that can regulate their tetrapyrrole content and composition in response to environmental signals. The present investigation analyses the interplay between light and dark regimes (continuous light, light-dark cycles (16:8) and continuous darkness) and aerobic, air-tight, and anaerobic environments (argon-enriched), on the relative composition of various pigments and growth attributes of Calothrix elenkenii as a prelude to exploiting C. elenkenii's bioindustrial potential as a source of pigments. Incubation in an anaerobic environment stimulated hormogonia formation and induced colouration/thickening of cells. Aerobically grown cultures of Calothrix, under continuous illumination produced the maximum amount of total phycobiliproteins and sugars, although chlorophyll accumulation and nitrogenase activity were highest in the light-dark environment. However, the beta-carotene content was observed to vary under anaerobic conditions with different light-dark regimes. This C. elenkenii strain can be a valuable source of pigments under optimized environmental conditions.     

Role of Toll-like receptors in antisense and siRNA [corrected

About 25 years ago, researchers first demonstrated that a short synthetic oligodeoxynucleotide, referred to as antisense, can inhibit replication of Rous sarcoma virus through hybridization to viral RNA. Since then, several hybridization-based oligonucleotide approaches have been developed to elucidate the functions of genes and their potential as therapeutic agents. Short-interfering (si) RNA is the most recent example. To effectively inhibit gene expression, an antisense or siRNA must be resistant to nucleases, be taken up efficiently by cells, hybridize efficiently with the target mRNA and activate selective degradation of the target mRNA or block its translation without causing undesirable side effects. However, both antisense and siRNA agents have been shown to exert non-target-related biological effects including immune stimulation. Do antisense and siRNA agents work as ligands for Toll-like receptors (TLRs), a family of pathogen-associated, molecular pattern recognition receptors?     

Self-stabilized CpG DNAs optimally activate human B cells and plasmacytoid dendritic cells

We recently showed that 5'-terminal secondary structures in CpG DNA affect activity significantly more than those at the 3'-end [Biochem. Biophys. Res. Commun. 306 (2003) 948]. The need for an accessible 5'-end of CpG DNA for activity suggested that the receptor reads the DNA sequence from this end. In continuation of these studies, we have designed immunomodulatory oligonucleotides (IMOs), consisting of a nine-mer stimulatory domain, containing a CpG motif and a hairpin-loop structure at the 3'-end, referred to as self-stabilized CpG DNAs. We studied the ability of self-stabilized CpG DNAs to stimulate human B-cell proliferation and interferon-alpha (IFN-alpha) secretion in plasmacytoid dendritic cell (pDC) culture assays. Self-stabilized CpG DNAs activated human B cells and induced plasmacytoid dendritic cells to secrete high levels of IFN-alpha. While both stimulatory and secondary structures in CpG DNAs were required for pDC activation, CpG motifs were sufficient to activate B cells. Interestingly, CpG motifs were not required for activity in the hairpin duplex region. Further modifications of the hairpin duplex region with a mixture of oligodeoxynucleotides and oligo-2'-O-methylribonucleotides in a heteroduplex formation permitted activation of both human B cells and pDCs.     

A genomic timescale for the origin of eukaryotes

Background  

Genomic sequence analyses have shown that horizontal gene transfer occurred during the origin of eukaryotes as a consequence of symbiosis. However, details of the timing and number of symbiotic events are unclear. A timescale for the early evolution of eukaryotes would help to better understand the relationship between these biological events and changes in Earth's environment, such as the rise in oxygen. We used refined methods of sequence alignment, site selection, and time estimation to address these questions with protein sequences from complete genomes of prokaryotes and eukaryotes.     

Evolutionary distance estimation under heterogeneous substitution pattern among lineages

Most of the sophisticated methods to estimate evolutionary divergence between DNA sequences assume that the two sequences have evolved with the same pattern of nucleotide substitution after their divergence from their most recent common ancestor (homogeneity assumption). If this assumption is violated, the evolutionary distance estimated will be biased, which may result in biased estimates of divergence times and substitution rates, and may lead to erroneous branching patterns in the inferred phylogenies. Here we present a simple modification for existing distance estimation methods to relax the assumption of the substitution pattern homogeneity among lineages when analyzing DNA and protein sequences. Results from computer simulations and empirical data analyses for human and mouse genes are presented to demonstrate that the proposed modification reduces the estimation bias considerably and that the modified method performs much better than the LogDet methods, which do not require the homogeneity assumption in estimating the number of substitutions per site. We also discuss the relationship of the substitution and mutation rate estimates when the substitution pattern is not the same in the lineages leading to the two sequences compared.     

Responses of isolated scale melanophores of a fresh water carp,Cirrhinus mrigala (Ham.) to zidovudine

Zidovudine (AZT) induced concentration related aggregation in C. mrigala melanophores. Denervated melanophores failed to respond to AZT. Specific and nonspecific alpha adrenoceptor antagonists completely blocked the responses of fish melanophores to AZT. Histamine and prostaglandin antagonists also inhibited aggregation of the melanophores induced by AZT. The results suggest that AZT may release a mixture of neurotransmitter like substances, which cause the aggregation of this fish melanophores.     

Baylis-Hillman reaction: convenient ascending syntheses and biological evaluation of acyclic deoxy monosaccharides as potential antimycobacterial agents

A series of acyclic deoxy carbohydrate derivatives from easily available carbohydrate enals 1, 2, 3 or 5 were prepared involving the Baylis-Hillman reaction. These newly formed carbohydrate based Baylis-Hillman adducts and their amino derivatives were evaluated for their antimycobacterial activity against Mycobacterium tuberculosis H(37)R(v). Among the compounds evaluated for their antimycobacterial activity, compound (10) showed the desired activity in the range of 3.125 microg/mL.     

Immunostimulatory properties of phosphorothioate CpG DNA containing both 3'-5'- and 2'-5'-internucleotide linkages

Synthetic oligodeoxyribonucleotides containing CpG-dinucleotides (CpG DNA) in specific sequence contexts activate the vertebrate immune system. We have examined the effect of 3′-deoxy-2′–5′-ribonucleoside (3′-deoxynucleoside) incorporation into CpG DNA on the immunostimulatory activity. Incorporation of 3′-deoxynucleosides results in the formation of 2′–5′-internucleotide linkages in an otherwise 3′–5′-linked CpG DNA. In studies, both in vitro and in vivo, CpG DNA containing unnatural 3′-deoxynucleoside either within the CpG-dinucleotide or adjacent to the CpG-dinucleotide failed to induce immunostimulatory activity, suggesting that the modification was not recognized by the receptors. Incorporation of the same modification distal to the CpG-dinucleotide in the 5′-flanking sequence potentiated the immunostimulatory activity of the CpG DNA. The same modification when incorporated in the 3′-flanking sequence had an insignificant effect on immunostimulatory activity of CpG DNA. Interestingly, substitution of a 3′-deoxynucleoside in the 5′-flanking sequence distal to the CpG-dinucleotide resulted in increased IL-6 and IL-10 secretion with similar levels of IL-12 compared with parent CpG DNA. The incorporation of the same modification in the 3′-flanking sequence resulted in lower IL-6 and IL-10 secretion with similar levels of IL-12 compared with parent CpG DNA. These results suggest that site-specific incorporation of 3′-deoxynucleotides in CpG DNA modulates immunostimulatory properties.     

Potent CpG oligonucleotides containing phosphodiester linkages: in vitro and in vivo immunostimulatory properties

Bacterial and synthetic DNAs, containing CpG dinucleotides in specific sequence contexts, activate the vertebrate immune system. Unlike phosphorothioate (PS) CpG DNAs, phosphodiester (PO) CpG DNAs require either palindromic sequences and/or poly(dG) sequences at the 3(')-end for activity. Here, we report 'PO-immunomers' having two PO-CpG DNA molecules joined through their 3(')-ends. These PO-imunomers permitted us, for the first time, to assess immunostimulatory properties of PO-CpG DNAs in vitro and in vivo without the need for palindromic and/or poly(dG) sequences. In medium containing 10% fetal bovine serum, PO-immunomers were more resistant than PO-CpG DNAs to nucleases. Compared to PS-CpG DNA in BALB/c and C3H/HeJ mice spleen cell culture assays, PO-immunomers showed increased IL-12 secretion and minimal amounts of IL-6 secretion. PO-immunomers activated NF-kappa B and induced cytokine secretion in J774 cell cultures. In addition, PO-immunomers showed antitumor activity in nude mice bearing human breast (MCF-7) and prostate (DU145) cancer xenografts.