Alkaloids of the leaves of Voacanga grandifolia

Besides vobtusine and vobtusine-lactone, deoxyvobtusine was isolated from the leaves of Voacanga grandifolia (Miq. Rolfe. Spectral and chemical evi     

Socioeconomic, demographic and environmental determinants of infant mortality in Nepal

The Nepal Fertility and Family Planning Survey of 1986 demonstrated that demographic variables, previous birth interval and survival of preceding child, still predominated as determinants of infant mortality, particularly in rural areas of Nepal. However, in urban Nepal, where the level of socioeconomic development is higher, an environmental variable, along with previous birth interval and survival of preceding child emerges as important in determining infant mortality. Separate policy measures for child survival prospects in rural and urban Nepal are suggested.     

A simple quantitative method for the estimation of free ecto-sulfhydryl groups of spermatozoa

A simple rapid quantitative method has been developed for the estimation of sperm ecto-SH groups on the basis of their high affinity binding to the mercurial: [203Hg]p-chloromercuriphenylsulfonic acid (PCMPS) used as a surface probe. The thiol reagent did not penetrate the sperm plasma membrane, as evidenced by the extremely rapid time course of the binding reaction and undetectable uptake of [203Hg]PCMPS by intact goat spermatozoa. The binding reaction was not due to contaminating broken or damaged cells, if any. The method consists of incubating of highly motile goat spermatozoa with PCMPS in a modified Ringer solution at 37 degrees C for 5 min, agglutination of the labelled cells with polyethyleneimine (100 micrograms/ml) and filtration and washing of the cell suspension through Whatman No. 1 filter discs under mild vacuum. The binding interaction is proportional to cell concentration, specific and saturable at 50 microM PCMPS. The method is capable of estimating free ecto-SH as low as 25 pmoles. Spermatozoa possess 286 +/- 61 pmoles of free ecto-SH groups/10(6) cells. Scatchard analysis showed the presence in goat spermatozoa of multiple classes of ecto-SH groups differing in their affinity for PCMPS.     

Probing binding mechanism of interleukin-6 and olokizumab: in silico design of potential lead antibodies for autoimmune and inflammatory diseases

Computer-aided antibody engineering has been successful in the design of new biologics for disease diagnosis and therapeutic interventions. Interleukin-6 (IL-6), a well-recognized drug target for various autoimmune and inflammatory diseases such as rheumatoid arthritis, multiple sclerosis, and psoriasis, was investigated in silico to design potential lead antibodies. Here, crystal structure of IL-6 along with monoclonal antibody olokizumab was explored to predict antigen–antibody (Ag???Ab)-interacting residues using DiscoTope, Paratome, and PyMOL. Tyr56, Tyr103 in heavy chain and Gly30, Ile31 in light chain of olokizumab were mutated with residues Ser, Thr, Tyr, Trp, and Phe. A set of 899 mutant macromolecules were designed, and binding affinity of these macromolecules to IL-6 was evaluated through Ag???Ab docking (ZDOCK, ClusPro, and Rosetta server), binding free-energy calculations using Molecular Mechanics/Poisson Boltzman Surface Area (MM/PBSA) method, and interaction energy estimation. In comparison to olokizumab, eight newly designed theoretical antibodies demonstrated better result in all assessments. Therefore, these newly designed macromolecules were proposed as potential lead antibodies to serve as a therapeutics option for IL-6-mediated diseases.     

c-kit protein, a transmembrane kinase: identification in tissues and characterization.

The proto-oncogene c-kit encodes a transmembrane kinase which is related to the receptors for colony-stimulating factor type 1 and platelet-derived growth factor, as well as to the immunoglobulin superfamily. Antibodies specific for the kinase domain of the P80 gag-kit protein of the Hardy-Zuckerman 4 feline sarcoma virus were prepared. These kit-specific antibodies were used to identify and characterize the c-kit protein in cat brain tissue. The c-kit protein product displays an autophosphorylating activity in immune complex kinase assays, and, in turn, this activity was used to identify the c-kit protein in different tissues. In cat brain, a single 145-kilodalton (kDa) glycoprotein was detected. Its N-linked carbohydrates were found to be sensitive to digestion with the endoglycosidases (neuraminidase, endoglycosidase F, and endoglycosidase H), indicating hybrid and/or complex and high-mannose structures. A partial purification of the c-kit protein was achieved by wheat germ agglutinin affinity chromatography, and the autophosphorylating activity of the partially purified c-kit protein was characterized and found to be specific for tyrosine. The kit antibodies cross-react with the murine c-kit protein product, and variant c-kit proteins in different mouse tissues were identified, with sizes of about 145 kDa (brain), 160 kDa (spleen), and 150 kDa (testis).     

A simple quantitative method of estimation of cell-intactness based on ethidium bromide fluorescence

A quantitative method based on fluorescence generated by the binding of ethidium bromide (EB) to DNA has been developed for estimation of the intactness of the plasma membrane of a mammalian cell type (goat epididymal spermatozoon). The method consists of mixing of sperm preparations with EB in a modified Ringer's solution followed by immediate measurement of fluorescence intensity at 365-580 nm (excitation-emission). The data were corrected for non-specific values of fluorescence due to intact cells only. The percentage of damaged cells in a sperm population was calculated by comparing the corrected fluorescence values of the cell preparations with those of the sonicated cells. The values of sperm intactness obtained by this method (99.5 +/- 0.3) compared well with those obtained by the widely used "marker enzyme" method (97 +/- 0.8) based on estimation of lactic dehydrogenase in the extracellular medium. The validity of this method has been confirmed by using cells of defined intactness i.e. preparations of vigorously forward-motile spermatozoa that showed nearly 100% intactness. The method can detect as low as 0.5% "leaky" or damaged cells in a cell preparation. The "EB-fluorescence" method is simpler and more rapid and reliable than the conventional "marker enzyme" method for estimation of cellular intactness.     

Sequence specificity of Z-DNA formation in oligonucleotides

The sequence specific requirement for B----Z transition in solution was examined in d(CGTGCGCACG), d(CGTACGTACG), d(ACGTACGT) in presence of various Z-inducing factors. Conformational studies show that inspite of the alternating nature of purines and pyrimidines, the aforementioned sequences do not undergo B----Z transition under the influence of NaCl, hexamine cobalt chloride and ethanol. A comparison with the crystal structures of an assorted array of purine and pyrimidine sequences show that the sequence requirement for B----Z transition is much more stringent in solution as compared to the solid state. The disruptive influence of AT base pairs in B to Z transition is discussed.