Hormonotoxins: the role of positive charge of lysine residue on the immunological,biological and cytotoxic properties of ovine lutropin-S-S-gelonin conjugates

Since the positive charge on the lysine residues plays an important role in the receptor recognition ability of oLH, the hormonotoxin has been synthesised with the use of 2-iminothiolane HC1 (2IT) and N-Succinimidyl-3-(2-pyridyldithio)-propionate (SPDP). The oLH activated with 2IT (oLH-10) was then mixed with SPDP activated gelonin (gelonin-30) in order to obtain a oLH-S-S-gelonin hormonotoxin. The conjugation mixture containing hormonotoxin was purified by gel-filtration chromatography according to the molecular weight and a complete physico-chemical, immunochemical and biochemical analysis were performed. The linkage occured through the -NH₂ groups of -subunit of oLH as judged from RP-HPLC analysis. A 11 (oLH:gelonin) molar ratio was obtained when determined with the use of several techniques. The hormonotoxins retained substantial receptor binding, steroidogenic activity and immunoreactivity. The competitive displacement analysis indicate that the binding occurs via the hormone part leaving the gelonin free which was probed with the gelonin antibodies. The presently described (C150A-02, C160A-02 and C170A-02) hormonotoxins exhibited higher receptor binding and toxicity to the target cells than the hormonotoxins prepared with the use of SPDP only. Therefore it is concluded that higher receptor binding and cytotoxicity may be due to the retention of positive charge on the lysine residues of oLH which was preserved during the conjugation process.Abbreviations BSA Bovine Serum Albumin - CMC Carboxy methyl Cellulose - DTT Dithiothreitol - DMEM Dulbeco's Modified Eagle's Medium - DTNB Ellman's reagent [5,5-dithio-bis-(2-nitrobenzoic acid)] - EDTA Ethylenediaminetetraacetic acid - FPLC Fast Protein Liquid Chromatography - FCA Freund's Complete Adjuvant - FCS Fetal Calf Serum - Gelonin-30 Gelonin modified by SPDP - GnRH Gonadotropin-Releasing Hormone - Gelonin-SPDP SPDP modified derivative of gelonin - HEPES (N-[2-hydroxyethyl] piperazine-N-[-2-ethanesulphonic acid]) - IFA Incomplete Freund's Adjuvant - 2IT 2-Iminothiolane - IODOGEN 1,3,4,6-tetrachloro 3,6-diphenylglycouril - oLH Ovine Luteinizing Hormone - oLH-SPDP SPDP modified derivative of oLH - oLH-10 oLH modified by 2IT - oLH2IT Molar ratio of oLH and 2IT - PDP 2-Pyridyl-dithiopropionate - PAP Pokeweed Antiviral Protein - RIP Ribosome Inactivating Protein - RP-HPLC Reverse-Phase High Performance Liquid Chromatography - RPMI Roswell Park Memorial Institute - RIA Radioimmunoassay - RRA Radioreceptor Assay - SPDP N-Succinimidyl-3(2-pyridyldithio)propionate - SDS-PAGE Sodium Dodecyl Sulphate-Polyacrylamide Gel Electrophoresis - TCA Trichloroacetic acid - TFA Trifluroacetic acid     

An H-2 haplotype possibly derived by crossing-over between the (A α A β ) duplex and the E βlocus

The B10.STA62 strain carries the H-2 ^w27 haplotype derived from a wild mouse captured in the vicinity of Ann Arbor, Michigan. Products of two class II loci composing this haplotype, A and A , are serologically, biochemically (by tryptic peptide mapping), and functionally indistinguishable from products controlled by the A ^b and A ^/b genes of the B10.A(5R) strain. In contrast, the polypeptide chain controlled by the third class II locus, E , is different from that controlled by the E ^/b gene. This E ^/w27 chain lacks an antigenic determinant present on the E^b molecule and carries determinants lacking on the E^b molecule, the E ^/b and E ^/w27 peptide maps differ in at least six peptides, and cytotoxic T cells specific for the E ^b chains do not react with B10.STA62 target cells. This great difference between the E ^/b and E ^/w27 chains suggests that the corresponding genes have not been derived from one another by a direct mutational conversion; instead, H-2 ^w27 appears to be a recombinant haplotype derived by crossing-over between the A A duplex and the E locus. This is the first recombinant discovered separating these class II loci.     

Survivability of parthenogenetic embryos following in vivo transfer in naturally synchronized Capra hircus

The present study was conducted to see the in vivo developmental potency of caprine parthenogenetic embryos generated in a modified way. The good quality caprine oocytes were matured in presence of cytochalasin B (CCB) and then activated by 7% ethanol followed by 2 mM 6-dimethyl amino purine (6-DMAP) and embryo development was recorded. Early stage parthenogenetic embryos (two to four cells) were surgically transferred in recipients (10). The pregnancy diagnosis was done by nonreturn to oestrus, ultrasonography (USG), and progesterone estimation. The levels of progesterone were above normal values (1 ng/ml) of pregnancy and fall below the level of pregnancy just before retuned to oestrus. Progesterone profile revealed that out of ten recipients (G1–G10), four goats (G1, G2, G3, and G5) returned to oestrus after 43?±?7.29 (Mean?±?SE) d of embryo transfer and six goats (G4, G6, G7, G8, G9, and G10) did not return to cycle even after 70 d of embryo transfer. In three recipients (G4, G5, and G6), the USG on day 40 revealed that there was fluid filled uterine body with solid fetus-like structure. These might be dead fetus and had started resorption. The progesterone profile also corroborated the assumption of pregnancy in these animals. Authors believe that this may be the first report on in vivo diploid parthenogenetic embryo development in caprine species.     

Anticonvulsant Effect of Asparagus racemosus Willd. in a Mouse Model of Catamenial Epilepsy

Neurochemical Research - Asparagus racemosus Willd. (Family Liliaceae), also known as female reproductive tonic, is traditionally used across the Sub-Himalayan region in Uttarakhand, India for...     

Transport and Metabolism of Sucrose versus Hexoses in Relation to Growth in Etiolated Pea Stem

Sucrose, supplied to detached pea (Pisum sativum L. var Alaska) epicotyls through cut bases, supported better growth of apical tissue than supplied glucose and/or fructose. The hexoses were converted mainly to sucrose in basal regions of the epicotyl but some moved as such through the epicotyl and accumulated at the apex (plumule) at a rate faster than sucrose. A greater proportion of the carbon derived from supplied hexoses than from sucrose was used for synthesis of ethanol-insoluble products throughout the epicotyl. By use of asymmetrically labeled sucrose, it was shown that neither hexose moiety was used preferentially for the synthesis of metabolites. Supplied sucrose moved as such only up to the region of cell elongation where it was hydrolyzed and completely equilibrated before moving into more apical regions. The results indicate that better growth with supplied sucrose than hexose could not have resulted from differential effects on cell division, more rapid uptake or transport of sucrose, enhanced wall synthesis, or cleavage by sucrose synthase. It is concluded that transported sucrose versus hexoses must undergo or evoke different reactions which affect growth in the region of cell elongation.     

Host chaperones are recruited in membrane-bound complexes by Plasmodium falciparum.

The ability of malarial parasite to deploy proteins at the surface of infected erythrocytes is well known. After their synthesis within the parasite, the cargo proteins are exported from the parasite and carried across the erythrocyte cytoplasm to be delivered at the erythrocyte surface. Our knowledge about the mechanisms involved in this complex trafficking path is limited. We have addressed the involvement of chaperones in traffic across erythrocyte cytoplasm. Our analyses of the chaperones available to the parasite indicated that none of the reported chaperones of the parasite origin are present in the erythrocyte cytoplasm. The chaperones of the host (Hsp70, Hsp90, Hop60), on the other hand, were readily detected in the erythrocyte cytosol. Hypotonic lysis and detergent solubilization experiments indicated that unlike their soluble nature in normal erythrocytes, host chaperones are recruited in membrane-bound, detergent-resistant complexes in infected cells. The association of host-Hsp70 with detergent-resistant complexes was ATP-dependent. Importantly, host chaperones could be detected in knob-enriched fractions and could be cross-linked to the knob subunit, PfHRP1, in a large complex at the surface of the infected erythrocytes. Our results implicate host chaperones in the assembly of parasite proteins such as knob subunits at the erythrocyte surface.     

Evaluation of Corrosion Resistance and Characterization of Ni-Cr Coated Structure Using Plasma Spray Coating for Marine Hulls

Plasmonics - Corrosion is a natural process which gradually destructs the materials with their environment by chemical or electrochemical means. Mechanized boat hull structures used in fishing were...     

The versatile role of glucose signalling in regulating growth,development and stress responses in plants

Journal of Plant Biochemistry and Biotechnology - Sugars as an energy source and a signalling molecule are indispensible for growth, development and stress responses in plants. Among sugars,...     

HY5-COP1: the central module of light signaling pathway

Journal of Plant Biochemistry and Biotechnology - Light acts catalytically to initiate a cascade of events to eventually regulate different aspects of plant development. The cascade of light signal...