Application of the multiple antigenic peptides (MAP) strategy to the production of prophormone convertases antibodies: Synthesis,characterization and use of 8-branched immunogenic peptides

Antiserum against an N-terminal sequence of murine prohormone convertase-1 (mPC1) incorporating the sequence immediatley following the junction between the putative pro-region and the active enzyme was obtained. This was accomplished using the multiple antigenic peptide (MAP) approach whereupon an 8-branched polylysine core to which are grafted multiple copies of a 16 amino acid peptide representing the N-terminal sequence of mPC1 (positions 84–99) was synthesized by solid-phase Fmoc chemistry. The ensuing peptide was purified and fully characterized by RP-HPLC, ¹H-NMR, amino acid composition, peptide sequencing and ion-spray mass spectrometry. The immunological properties of the resulting antibodies in detecting recombinant PC1 in both crude and purified preparations were compared with antibodies raised against a similar N-terminal segment of PC1 but using the conventioanl method of peptide–carrier protein conjugation and also developed against a C-terminal fusion protein of PC1. Our data indicate that the MAP antibody was as efficient as both the amino and carboxy-terminal antibodies in qualitative as well as quantitative analysis of PC1 encoded protein by radioimmunoassay. Following an identical approach, antibodies against other prohormone convertases like furin, PC5/6 and PACE4 were also developed and subsequently applied to a number of biochemical and immunological studies. In each case, the ease of preparation and high immunogenicity of the MAP approach were confirmed and reside in the simplicity and rapidity with which a potent and useful antiserum is obtained.     

Metabolic changes during rooting in stem cuttings of five mangrove species

Vegetative propagation through rooting in stem cuttings in five tree mangroves namely Bruguiera parviflora, Cynometra iripa, Excoecaria agallocha, Heritiera fomes, and Thespesia populnea using IAA, IBA and NAA was reported. Spectacular increase in the root number was noted in the cuttings of H. fomes and C. iripa treated together with IBA (5000 ppm) and NAA (2500 ppm). The highest number of roots was obtained with IBA (2500 ppm) and NAA (500 ppm) in E. agallocha. B. parviflora and T. populnea responded better to IAA and IBA treatment. The species specific variation in the rooting response to exogenous application of auxins was reflected in the metabolic changes during initiation and development of roots in cuttings. Biochemical analysis showed increase of reducing sugar in the above-girdled tissues at initiation as well as subsequent development of roots which was further enhanced by the use of auxins. Decreases in the total sugar, total carbohydrate and polyphenols and increase in total nitrogen were recorded in the girdled tissues and the high C/N ratio at the initial stage helped in initiation of roots in all the species. Interaction of IBA and NAA promoted starch hydrolysis better than IAA and IBA during root development and subsequently reduced the C/N ratio and increased the protein-nitrogen activity during root development which suggest the auxin influenced mobilization of nitrogen to the rooting zone.Abbreviations IAA Indole-3-acetic acid - IBA Indole-butyric acid - NAA A-naphthalene acetic acid     

Melovinone,an open chain analogue of melochinone from Melochia tomentosa

Melovinone, a new alkaloid isolated from the roots of Melochia tomentosa has been characterized as 3,7,8-trimethoxy-2-methyl-5(5′-phenylpentyl)-4-quinolinone.     

A Geometry-Based Multiple Testing Correction for Contingency Tables by Truncated Normal Distribution

Statistics in Biosciences - Inference procedure is a critical step of experimental researches to draw scientific conclusions especially in multiple testing. The false positive rate increases unless...     

New Computational Approach for Peptide Vaccine Design Against SARS-COV-2

International Journal of Peptide Research and Therapeutics - The design for vaccines using in silico analysis of genomic data of different viruses has taken many different paths, but lack of any...     

In vitro morphogenesis in Selaginella microphylla (Kunth.) Spring

Selaginella, an extant genus of primitive vascular plants, has survived over 400 million years of evolution. In vitro morphogenesis in Selaginella microphylla is considered for the first time to establish a well-documented aseptic culture on half- strength Murashige and Skoog’s basal medium with 2ip (4.92–49.21 μM), or Kn (4.65–46.47 μM) or GA₃ (2.89–28.90 μM) for shoot multiplication, and with different concentrations of IBA (4.9–49 μm) to initiate root cultures. GA₃ was instrumental for shoot multiplication as well as induction of reproductive structures in each and every leaf axil. On the other hand, it is observed that IBA alone in S. microphylla can act as signal molecules for induction of enormous numbers of root masses from a few existing roots. An interesting pattern of re-differentiation has also been observed where apical portions of large numbers of roots were converted to green shoot apical meristems. Further differentiation produced tiny green shoots. Distinct bipolarity was noted in shoots when they were isolated from root masses and appeared as embryo-like structures. Chromosome analysis from in vitro sporophytic plants revealed 2n = 16 chromosomes, indicating chromosomal stability. The interesting in vitro pattern of morphogenesis obtained in S. microphylla may provide new insights into totipotency of plants.     

Emerging facets of plastid division regulation

Plastids are complex organelles that are integrated into the plant host cell where they differentiate and divide in tune with plant differentiation and development. In line with their prokaryotic origin, plastid division involves both evolutionary conserved proteins and proteins of eukaryotic origin where the host has acquired control over the process. The plastid division apparatus is spatially separated between the stromal and the cytosolic space but where clear coordination mechanisms exist between the two machineries. Our knowledge of the plastid division process has increased dramatically during the past decade and recent findings have not only shed light on plastid division enzymology and the formation of plastid division complexes but also on the integration of the division process into a multicellular context. This review summarises our current knowledge of plastid division with an emphasis on biochemical features, the functional assembly of protein complexes and regulatory features of the overall process.     

Fatty acid-induced angiogenesis in first trimester placental trophoblast cells: Possible roles of cellular fatty acid-binding proteins

Angiogenesis is involved in the growth of new blood vessels from the existing one. Consequently, angiogenesis plays an indispensable role in tissue growth and repair including early placentation processes. Besides angiogenic growth factors (vascular endothelial growth factor (VEGF), angiopoietin-like 4 (ANGPTL4), placental growth factor (PlGF), platelet derived growth factor (PDGF), fibroblast growth factors (FGF)), dietary fatty acids (c>16) also directly or indirectly modulate angiogenic processes in tumors and other cell systems. Usually n − 3 fatty acids inhibit whereas n − 6 fatty acids stimulate angiogenesis in tumors and other cells. Contrary to this, docosahexaenoic acid, 22:6n − 3 (DHA) and other fatty acids including conjugated linoleic acid stimulate angiogenesis in placental first trimester cells. In addition to the stimulation of expression of major angiogenic factors such as VEGF and ANGPTL4, fatty acids also stimulate expression of intracellular fatty acid-binding proteins (FABPs) FABP-4 and FABP-3 those are known to directly modulate angiogenesis. Emerging data indicate that FABPs may be involved in the angiogenesis process. This paper reviews the fatty acid mediated angiogenesis process and the involvement of their binding proteins in these processes.     

Clostridium perfringens epsilon toxin H149A mutant as a platform for receptor binding studies

Clostridium perfringens epsilon toxin (Etx) is a pore‐forming toxin responsible for a severe and rapidly fatal enterotoxemia of ruminants. The toxin is classified as a category B bioterrorism agent by the U.S. Government Centres for Disease Control and Prevention (CDC), making work with recombinant toxin difficult. To reduce the hazard posed by work with recombinant Etx, we have used a variant of Etx that contains a H149A mutation (Etx‐H149A), previously reported to have reduced, but not abolished, toxicity. The three‐dimensional structure of H149A prototoxin shows that the H149A mutation in domain III does not affect organisation of the putative receptor binding loops in domain I of the toxin. Surface exposed tyrosine residues in domain I of Etx‐H149A (Y16, Y20, Y29, Y30, Y36 and Y196) were mutated to alanine and mutants Y30A and Y196A showed significantly reduced binding to MDCK.2 cells relative to Etx‐H149A that correlated with their reduced cytotoxic activity. Thus, our study confirms the role of surface exposed tyrosine residues in domain I of Etx in binding to MDCK cells and the suitability of Etx‐H149A for further receptor binding studies. In contrast, binding of all of the tyrosine mutants to ACHN cells was similar to that of Etx‐H149A, suggesting that Etx can recognise different cell surface receptors. In support of this, the crystal structure of Etx‐H149A identified a glycan (β‐octyl‐glucoside) binding site in domain III of Etx‐H149A, which may be a second receptor binding site. These findings have important implications for developing strategies designed to neutralise toxin activity.     

Differential expression and functions of neuronal and glial neurofascin isoforms and splice variants during PNS development

The cell adhesion molecule neurofascin (NF) has a major neuronal isoform (NF186) containing a mucin-like domain followed by a fifth fibronectin type III repeat while these domains are absent from glial NF155. Neuronal NF isoforms lacking one or both of these domains are expressed transiently in embryonic dorsal root ganglia (DRG). These two domains are co-expressed in mature NF186, which peaks in expression prior to birth and then persists almost exclusively at nodes of Ranvier on myelinated axons. In contrast, glial NF155 is only detected postnatally with the onset of myelination. All these forms of NF bound homophilically and to Schwann cells but only the mature NF186 isoform inhibits cell adhesion, and this activity may be important in formation of the node of Ranvier. Schwann cells deficient in NF155 myelinated DRG axons in a delayed manner and they showed significantly decreased clustering of both NF and Caspr in regions where paranodes normally form. The combined results suggest that NF186 is expressed prenatally on DRG neurons and it may modulate their adhesive interactions with Schwann cells, which express NF155 postnatally and require it for development of axon-glial paranodal junctions.