Polycations. 19. The synthesis of symmetrical dicationic lipids with internal dimethylazonia functionalities separated by a spacer unit and pendant chains

Herein is reported the preparation of several series of symmetrical polyammonium salts that serve as cationic lipids or precursors thereof, and are structurally based on several series of parent diamines where dimethylazonia functionalities are present, separated by a central structural unit, and pendant terminal chains. The resultant materials are of significant interest for a variety of purposes, such as serving as antihydrophobic species and as transfectins, the details of which are provided in separate reports. Attempts to effect selective alkylation to provide the corresponding unsymmetrical cationic lipids were without success, always leading to relatively useless mixtures of products.     

Supramolecular zinc(II)salphen motifs: Reversible dimerization and templated dimeric structures

The formation of a dimeric structure of a nonsymmetric Zn(II)salphen complex is reported. The X-ray molecular structure show the formation of an oxygen-bridged species (2). In addition to this structure, a pyridine-ligated complex and an 1:2 dabco/Zn(II)salphen supramolecular assembly (dabco = diazabicyclo[2.2.2]octane) are presented. Their coordination behavior has been studied and can be correlated with the substitution pattern of the salphen ligand and the donor-strength of the involved axial ligands. The Zn(II)salphen building blocks bind in a cooperative fashion to the dabco template, the second unit being bound 4 times more strongly.     

Photooxidative degradation of beer bittering principles: product analysis with respect to lightstruck flavour formation.

Isohumulones, the main bittering agents in beer, are decomposed by light-induced reactions, thereby leading to radical precursors on the pathway to lightstruck flavour formation. Excited flavins, formed on visible-light irradiation, readily interact with isohumulones, as well as with reduced and oxidized derivatives thereof. From identification of both volatile and non-volatile reaction products thus formed, feasible degradation mechanisms are proposed.     

An ecological interpretation of the difference in leaf anatomy and its plasticity in contrasting tree species in orange kloof,table mountain,South Africa

Leaf anatomy and morphology were studied in 11 tree species growing in an undisturbed forest and the adjoining fynbos for over 50 years. Functional anatomical results suggest that the forest and the fynbos are ecologically distinct. Moreover, leaf anatomy suggests that the foliage is primarily adapted for photosynthesis rather than for control of transpirational water loss. Forest precursor tree species and scrub species exhibit xeromorphy in the fynbos whereas they exhibit mesomorphic features inside the forest. The wide-ranging species, such as Olea capensis subsp. capensis, simulated the response of the forest precursors, with the cuticle being phenotypically plastic between the forest and the fynbos but not between the stream and non-stream habitats. Finally, the forest precursors, the scrub species, and the wide-ranging taxa seem to have anatomical characters which can be modified in the fynbos and therefore allow its colonization by a variety of different species.     

Degradation of wild-type vasopressin precursor and pathogenic mutants by the proteasome

Mutations in the gene encoding the antidiuretic hormone arginine vasopressin cause autosomal dominant neurogenic diabetes insipidus. Autoptic data in affected individuals suggest that the neurons expressing mutant vasopressin undergo selective degeneration. Expression studies have shown that the mutants are retained in the endoplasmic reticulum, but how this trafficking defect is linked to neurotoxicity is unknown. One possibility is that unsecreted mutant precursors, or degradation products thereof, are cytotoxic. We therefore investigated the fate of endoplasmic reticulum-retained pathogenic mutants. Our data show that the mutants are retrotranslocated to the cytosol and degraded by the proteasome. In the presence of proteasomal inhibitors, three distinct un- or deglycosylated cytosolic species of vasopressin precursors were stabilized: pre-pro-vasopressin, pro-vasopressin, and an N-terminally truncated form. In addition to the retrotranslocated forms, a fraction of the newly synthesized precursor was not translocated, but was synthesized into the cytosol due to inefficient function of the vasopressin signal peptide. As a result, cytosolic pre-pro-vasopressin and its degradation product were also recovered when wild-type vasopressin was expressed. Cytosolic forms of vasopressin might trigger cytotoxicity in vivo, as has been proposed in the case of prion protein, which also contains an inefficient N-terminal signal peptide.     

Multifunctional peptides encrypted in milk proteins

Many bioactivities of milk are latent in that they are inactive within the protein sequence, requiring enzymatic proteolysis for release of bioactive peptides from milk proteins precursors. Bioactivities of peptides encrypted in major milk proteins are latent until released and activated, e.g. during gastrointestinal digestion or food processing. Bioactive peptides can be produced in vivo following intake of milk proteins, and the proteolytic system of bacterial species used in the production of fermented milk products and cheese can contribute to the liberation of bioactive peptides or precursors thereof. Activated peptides are potential modulators of various regulatory processes in the living system: immunomodulatory peptides stimulate the activities of cells of the immune system and several cytomodulatory peptides inhibit cancer cell growth, antimicrobial peptides kill sensitive microorganisms, angiotensin-I-converting enzyme (ACE)-inhibitory peptides exert an hypotensive effect, opioid peptides are opioid receptor ligands which can modulate absorption processes in the intestinal tract, mineral binding peptides may function as carriers for different minerals, especially calcium. Many milk-derived peptides reveal multifunctional properties, i.e. specific peptide sequences having two or more different biological activities have been reported. Milk protein-derived bioactive peptides are claimed to be health enhancing components that can be used to reduce the risk of disease or to enhance a certain physiological function.     

Pigment pattern evolution by differential deployment of neural crest and post-embryonic melanophore lineages in Danio fishes

Latent precursors or stem cells of neural crest origin are present in a variety of post-embryonic tissues. Although these cells are of biomedical interest for roles in human health and disease, their potential evolutionary significance has been underappreciated. As a first step towards elucidating the contributions of such cells to the evolution of vertebrate form, we investigated the relative roles of neural crest cells and post-embryonic latent precursors during the evolutionary diversification of adult pigment patterns in Danio fishes. These pigment patterns result from the numbers and arrangements of embryonic melanophores that are derived from embryonic neural crest cells, as well as from post-embryonic metamorphic melanophores that are derived from latent precursors of presumptive neural crest origin. In the zebrafish D. rerio, a pattern of melanophore stripes arises during the larval-to-adult transformation by the recruitment of metamorphic melanophores from latent precursors. Using a comparative approach in the context of new phylogenetic data, we show that adult pigment patterns in five additional species also arise from metamorphic melanophores, identifying this as an ancestral mode of adult pigment pattern development. By contrast, superficially similar adult stripes of D. nigrofasciatus (a sister species to D. rerio) arise by the reorganization of melanophores that differentiated at embryonic stages, with a diminished contribution from metamorphic melanophores. Genetic mosaic and molecular marker analyses reveal evolutionary changes that are extrinsic to D. nigrofasciatus melanophore lineages, including a dramatic reduction of metamorphic melanophore precursors. Finally, interspecific complementation tests identify a candidate genetic pathway for contributing to the evolutionary reduction in metamorphic melanophores and the increased contribution of early larval melanophores to D. nigrofasciatus adult pigment pattern development. These results demonstrate an important role for latent precursors in the diversification of pigment patterns across danios. More generally, differences in the deployment of post-embryonic neural crest-derived stem cells or their specified progeny may contribute substantially to the evolutionary diversification of adult form in vertebrates, particularly in species that undergo a metamorphosis.     

Structure and function of plant aspartic proteinases.

Aspartic proteinases of the A1 family are widely distributed among plant species and have been purified from a variety of tissues. They are most active at acidic pH, are specifically inhibited by pepstatin A and contain two aspartic residues indispensible for catalytic activity. The three-dimensional structure of two plant aspartic proteinases has been determined, sharing significant structural similarity with other known structures of mammalian aspartic proteinases. With a few exceptions, the majority of plant aspartic proteinases identified so far are synthesized with a prepro-domain and subsequently converted to mature two-chain enzymes. A characteristic feature of the majority of plant aspartic proteinase precursors is the presence of an extra protein domain of about 100 amino acids known as the plant-specific insert, which is highly similar both in sequence and structure to saposin-like proteins. This insert is usually removed during processing and is absent from the mature form of the enzyme. Its functions are still unclear but a role in the vacuolar targeting of the precursors has been proposed. The biological role of plant aspartic proteinases is also not completely established. Nevertheless, their involvement in protein processing or degradation under different conditions and in different stages of plant development suggests some functional specialization. Based on the recent findings on the diversity of A1 family members in Arabidopsis thaliana, new questions concerning novel structure-function relationships among plant aspartic proteinases are now starting to be addressed.     

Regulation of differentiation in normal and transformed erythroid cells.

Studies are described employing two erythropoietic systems to elucidate regulatory mechanisms that control both normal erythropoiesis and erythroid differentiation of transformed hemopoietic precursors. Evidence is provided suggesting that normal erythroid cell precursors require erythropoietin as a growth factor that regulates the number of precursors capable of differentiating. Murine erythroleukemia cells proliferate without need of erythropoietin; they show a variable, generally low, rate of spontaneous differentiation and a brisk rate of erythropoiesis in response to a variety of chemical agents. Present studies suggest that these chemical inducers initiate a series of events including cell surface related changes, alterations in cell cycle kinetics, and modifications of chromatin and DNA structure which result in the irreversible commitment of these leukemia cells to erythroid differentiation and the synthesis of red-cell-specific products.     

Multicomponent synthesis of novel amino acid-nucleobase chimeras: a versatile approach to PNA-monomers

This paper describes a multicomponent approach to novel totally protected precursors of PNA-monomers via Ugi 4CC. The obtained bisamides are converted into several partially protected PNA-monomers or derivatives thereof using three different procedures. Methods for hydrolysis are shown to be dependent on the nature of the isocyano component required for Ugi 4CC. Several novel monomers suitable for oligomer synthesis are prepared demonstrating the high versatility of the reaction sequence.     

Porcine prion protein amyloid

ABSTRACT

Mammalian prions are composed of misfolded aggregated prion protein (PrP) with amyloid-like features. Prions are zoonotic disease agents that infect a wide variety of mammalian species including humans. Mammals and by-products thereof which are frequently encountered in daily life are most important for human health. It is established that bovine prions (BSE) can infect humans while there is no such evidence for any other prion susceptible species in the human food chain (sheep, goat, elk, deer) and largely prion resistant species (pig) or susceptible and resistant pets (cat and dogs, respectively). PrPs from these species have been characterized using biochemistry, biophysics and neurobiology. Recently we studied PrPs from several mammals in vitro and found evidence for generic amyloidogenicity as well as cross-seeding fibril formation activity of all PrPs on the human PrP sequence regardless if the original species was resistant or susceptible to prion disease. Porcine PrP amyloidogenicity was among the studied. Experimentally inoculated pigs as well as transgenic mouse lines overexpressing porcine PrP have, in the past, been used to investigate the possibility of prion transmission in pigs. The pig is a species with extraordinarily wide use within human daily life with over a billion pigs harvested for human consumption each year. Here we discuss the possibility that the largely prion disease resistant pig can be a clinically silent carrier of replicating prions.     

Preparation of synthetic tRNA precursors with tRNA nucleotidyltransferase.

Rabbit liver tRNA nucleotidyltransferase can be used to substitute nucleotides within the -C-C-A sequence of tRNA or to add nucleotides following this sequence. These anomolous reactions of the enzyme have been used to prepare radioactively-labeled synthetic tRNA precursors which mimic the structure of the natural precursors. Under appropriate conditions synthetic precursors of defined structure can be made. In this paper we describe the synthesis of tRNA-C-[14C]U and tRNA-C-C-A-[14C]C-C, which are representative of tRNA precursors containing altered residues within the -C-C-A sequence or with extra residues following the normal 3'terminus. A variety of other possible precursors can also be prepared. These synthetic tRNA precursors have already proved useful for isolation of possible tRNA processing nucleases.     

Transfer RNA splicing in Saccharomyces cerevisiae. Secondary and tertiary structures of the substrates

Secondary and tertiary structures of four yeast tRNA precursors that contain introns have been elucidated using limited digestion with a variety of single-strand- and double-strand-specific nucleases. The pre-tRNAs, representing the variety of intron sizes and potential structures, were: pre-tRNALeuCAA, pre-tRNALeuUAG, pre-tRNAIleUAU, and pre-tRNAPro-4UGG. Conventional tRNA cloverleaf structure is maintained in these precursors except that the anticodon loop is interrupted by the intron. The intron contains a sequence which is complementary to a portion of the anticodon loop and allows the formation of a double helix often extending the anticodon stem. The 5' and 3' splicing cleavage sites are located at either end of this helix and are single-stranded. The intron is the most sensitive region to nuclease cleavage, suggesting that it is on the surface of the molecule and available for interaction with the splicing endonuclease. Absence of Mg2+ or spermidine renders the dihydrouridine and T psi C loops of these precursors highly sensitive to nuclease digestion. These ionic effects mimic those observed for tRNAPhe and suggest that the tRNA portion of these precursors has native tRNA structure. We propose consensus secondary and tertiary structures which may be of significance to eventual understanding of the mechanism of yeast tRNA splicing.     

Biosynthesis of amaranthin in Celosia plumosa

Seedlings of a yellow betaxanthin-producing variety of Celosia plumosa when fed with appropriate precursors are capable of synthesizing the red-violet pigment normally present in red varieties of the same species, namely amaranthin. Synthesis of amaranthin occurs in seedlings following administration of betanidin and betanin but much greater accumulation was observed after feeding cycloDOPA and its 5-O-β-d-glucoside. Possible pathways in the biosynthesis of amaranthin are discussed.     

Membrane proteins specified by herpes simplex viruses. I. Identification of four glycoprotein precursors and their products in type 1-infected cells.

Polypeptide precursors to the major glycoproteins specified by herpes simplex virus type 1 were identified in immunoprecipitation experiments using antisera that reacted specifically with the viral glycoproteins and their precursors. The results demonstrate that the major glycosylated proteins detected in infected cells are derived from four antigenically distinct polypeptides. Three of these polypeptides become glycosylated in two discrete stages, yielding partially glycosylated intermediates and fully glycosylated products. The final products are the predominant species detected in cytoplasmic virions and in plasma membranes. The fourth polypeptide precursor appears to acquire very little carbohydrate and differs in several respects from the other three precursors.     

Multiple bombesin-like peptides with opposite functions from skin of Odorrana grahami

Bombesin-like peptides (BLPs) are a family of neuroendocrinic peptides that mediate a variety of biological activities. Three mature BLPs from the skin secretions of the frog Odorrana grahami were purified. Several bombesin-like peptide cDNA sequences encoding precursors of BLPs were identified from the skin cDNA library of O. grahami. This is the maximal diversity of BLPs ever found in animals. Five mature BLPs (B1-B5) based on the amino acid sequences derived from the cDNA cloning were synthesized. In the in vitro myotropic contraction experiment, all synthesized BLPs displayed a stimulating effect toward rat stomach strips, except B4 and B5 which showed the opposite effect, suggesting that certain BLPs may act as antagonists of bombesin receptors while most other BLPs act as agonists. This finding will facilitate the finding of novel bombesin receptors and novel ligands of bombesin receptors. The diversity of amphibian BLPs and their precursors were also analyzed and results suggest that amphibian BLPs and corresponding precursors of various sizes and processing patterns can be used as markers of taxonomic and molecular phylogenetics. The remarkable similarity of preproregions gives rise to very different BLPs and 3'-terminal regions in distantly related frog species, suggesting that the corresponding genes form a multigene family originating from a common ancestor. The diversification of BLP loci could thus be part of an evolutionary strategy developed by amphibian species as a result of shifts to novel ecological niches when environmental factors change rapidly.     

The Molecular Species of Diacylglycerol Precursors Used in Monogalactosyldiacylglycerol Biosynthesis in the Leaves of a Number of Higher Plant Species

The molecular species composition of diacylglycerol precursorsof monogalactosyldiacyl-glycerol (MGDG) from a variety of agriculturallyimportant plant species have been determined using in vivo ¹⁴C-tracertechniques. Analysis of 16:3-plants shows distinct similaritiesbetween different species and the data support the theory oftwo separate pathways for the biosynthesis of 16C/18C and 18C/18Cmolecular species of MGDG. The analyses of 18:3-plants confirmthat the DAG precursors of MGDG in these species are highlyunsaturated containing mainly linoleic (18:2) and linolenicacid (18:3). They also show some important variations betweenspecies. (Received February 19, 1988; Accepted May 11, 1988)     

Postirradiation dynamics of T-lymphocyte precursors and thymus regeneration in mice

Irradiation of CBA mice elicits a pronounced response of intrathymus and bone marrow precursors of T-lymphocytes (PTL) which is manifested by an increase in a relative content of PTL in the thymus and their transfer from bone marrow to thymus. The estimate of the regenerative potency of the thymus most adequately reflects the cellular events leading to regeneration thereof. The differences in the thymus regeneration dynamics between CBA and AKR mice are associated with different radiation response of the intrathymic SC-1-PTL in mice of these strains.