A comparative study of cyclization strategies applied to the synthesis of head-to-tail cyclic analogs of a viral epitope.

A family of head-to-tail cyclic peptide models of the antigenic site A (G-H loop of viral protein 1) of foot-and-mouth disease virus has been designed on the basis of the three-dimensional structure adopted by the linear peptide YTASARGDLAHLTTT upon binding to neutralizing monoclonal antibodies. Three different methods of cyclization have been examined to access the peptides. Solution cyclization of a minimally protected linear precursor provided the expected products but required several purification steps that lowered the yields to approximately 10%. The two other approaches relied on side-chain anchoring of the peptide through the Asp residue and cyclization on the solid phase. A synthetic scheme combining Fmoc, tBu and OAI protections was practicable but inefficient when scaled-up. The combination of Boc, Bzl and OFm protections was more promising, but suffered from high epimerization during the initial esterification of Boc-Asp-OFm to benzyl alcohol-type resins. This problem was solved by performing the esterification via the cesium salt of Boc-Asp-OFm. With this improvement, the Boc/Bzl/OFm has become the method of choice for the preparation of cyclic head-to-tail peptides in satisfactory yields and with minimal purification.     

Leghemoglobin in Lupin Plants (Lupinus albus cv Multolupa)

Leghemoglobin was localized by immunogold techniques in nodules of Lupinus albus cv Multolupa inoculated with Bradyrhizobium sp. (Lupinus) strain ISLU 16. The protein localization was performed in nodules embedded in Spurr's and Araldite epoxy resins and Lowycryl K4M. A very good preservation of both the ultrastructure and antigenicity was obtained with the tissues embedded in Araldite following glutaraldehyde fixation and unpostfixed in osmium tetroxide. Lupin leghemoglobin is a stable and abundant protein which allows a conventional method to be safely used for localization of leghemoglobin. Labeling of leghemoglobin was specifically confined to the cytosol matrix and nuclei. Gold particles were never observed in the peribacteroidal spaces nor in the cytoplasmic organelles of the infected cells. Decrease of leghemoglobin was observed when the plants were grown with 10.7 micromolar and 21.4 micromolar of nitrate.     

Distribution of epiplanktonic Chaetognatha along a transect in the Indian Ocean

Chaetognaths were identified and counted in 23 samples collectedin the west Indian Ocean along a transect from the Gulf of Adento the Cape of Good Hope, in February–March 1967. Althoughthe hydrographic front located at 5°S does not seem to representa barrier for the distribution of the majority of the species,some of them are in a preferential area south or north of thefront. The principal component analysis indicates the locationswhere the more intense overlapping of species takes place. Itis important to mention the occurrence of S.pseudoserratodentatain the proximity of Madagascar Island.     

Immunoregulatory effects of a suppressor factor from healthy pregnant women's lymphocytes after progesterone induction

Progesterone-treated pregnancy lymphocytes release an immunologic blocking factor. The mode of action of this substance was investigated. The supernatant of progesterone-treated pregnancy lymphocytes was highly suppressive of natural cytotoxicity toward human embryonic fibroblast target cells as well as of natural killer cell activity. The effect was not observed when progesterone induction was performed in the presence of RU 486, a progesterone receptor blocking agent. The factor was able to inhibit mixed lymphocyte reactions (MLRs), and transfer coculture experiments revealed that this effect was dependent on major histocompatibility complex nonspecific, nonrestricted suppressor T cells. The activation/expansion of suppressor inducer and suppressor effector T cells was further proved by fluorescence-activated cell sorter analysis of the populations from MLRs cultured in the presence of the inhibitory factor. These changes were not observed with MLRs performed in the presence of supernatants from progesterone + RU 486-treated peripheral blood lymphocytes. The inhibitory material, on the other hand, did not affect either production or function of IL-2. We conclude that in the presence of high local concentrations of progesterone, a suppressive pathway dependent on specific progesterone-CD8+ lymphocyte interaction might be established. This mechanism might play an important role in the maintenance of pregnancy.     

Binding and action of cecropin and cecropin analogues: antibacterial peptides from insects

The mechanism of action of cecropin was studied by using liposomes as a model system. The bilayer was efficiently destroyed if the liposome net charge was zero or negative. Cecropin analogues with an impaired N-terminal helix had reduced membrane disrupting abilities that correlate with their lower antibacterial activity. The reduced bactericidal activity of the analogues was rationalized in terms of reduced binding to bacteria. The stoichiometry of cecropin killing of bacteria suggests that amounts of cecropin sufficient to form a monolayer strongly modify the bacterial membrane. Although some bacteria were resistant to cecropin they did bind large amounts in a non-productive manner. In contrast, mammalian erythrocytes achieve resistance by avoiding the binding of cecropin.     

Interactions of ubiquinones with membrane models

The effect of the insertion of coenzyme Q10 and some of its shorter chain homologues in membrane models (Reverse Micelles, Small Unilamellar Vesicles and Liposomes) has been studied by NMR and IR spectroscopies. By 1H-NMR we have found that the stretched conformation of the isoprenoid side-chain observed in solution is maintained in membrane models. Interaction between the quinonoid moiety of the Q's and the phosphatidic groups of the phospholipids has been evidenced by 31P-NMR. A large effect of this interaction on the water microdynamics and distribution around the charged groups of the phospholipids has been observed by measurements of 1H and 2H relaxation times and by infrared spectra. The 13C-NMR spectra of the backbone of the acyl chains of phospholipids does not seem to be influenced to a noticeable extent by the presence of the Q's.     

Supramolecular organization of lysophosphatidylcholine-packaged Gramicidin A

Heat derived gramicidin A'/L-alpha-lysophosphatidylcholine complexes were separated on a sucrose gradient to form two fractions: Fraction A which had an approximately constant Gramicidin A' to phospholipid ratio of 8 to 10 lipid molecules per Gramicidin A' molecule and Fraction B which had a larger but variable ratio. Fluorescence and circular dichroism studies confirmed Fraction A to be a lipid-incorporated channel state. Electron microscopic studies, using uranyl acetate negative staining, showed fraction A to be a membranous state with the formation of bilayer vesicles, that is, the interaction of peptide and phospholipid micelles causes the lipid to reorganize into a bilayer structure. Freeze-fracture replicas of the channel incorporated state demonstrated the presence of a supramolecular organization of particles exhibiting a tendency to form rows with a 50-60 A periodicity along the row and with 70-80 A distance between rows. An idealized working model for the incorporated state is presented.