Phosphatidic acid, phosphatidylinositol, phosphatidylserine and cardiolipin in the course of early embryonic development. Fatty acid composition and content in whole toad embryos and in mitochondrial fractions

The fatty acid composition and content of phosphatidylinositol, phosphatidylserine and phosphatidic acid have been studied during the early development of toad embryos. Acidic phospholipids have been analyzed in whole oocytes and embryos and in the following subcellular fractions: yolk platelets, mitochondria and microsomes. Also cardiolipin, a mitochondrial phospholipid, has been analyzed. Gastrula stage embryos have shown, mainly in the mitochondrial fraction, an increase in the content of phosphatidic acid, phosphatidylserine and phosphatidylinositol with respect to unfertilized oocytes. Changes in the distribution of acyl groups of phosphatidic acid have been detected when different subcellular fractions are compared. On the other hand, the phosphatidylserine composition remains unmodified. Arachidonate and stearate are the principal components of phosphatidylinositol. Cardiolipin shows the same composition up to gastrulation and linoleate comprises about 50% of the total acyl groups.     

1,6-diamino-2,5-anhydro-1,6-dideoxy-l-iditol and some derivatives thereof

1,6-Diamino-2,5-anhydro-1,6-dideoxy-l-iditol (31) and its derivatives were synthesized, starting from 2,4-O-benzylidene-1,6-di-O-tosyl-d-glucitol. The 1,6-bis-(acetamido)-l-talo epoxide was readily hydrolyzed to the corresponding l-iditol derivative under anchimeric assistance of the 1-acetamido group. On treatment with formaldehyde-formic acid, diamine 31 gave a tricyclic, 1,4:3,6-bis(N,O-methylene) derivative which was stable under acidic conditions but, according to ¹³C-n.m.r. spectroscopy, was readily hydrolyzed to an equilibrium mixture in neutral, aqueous solution. The corresponding 1,6-bis(dimethylamino) derivative could be obtained by reducing this equilibrium mixture with borohydride. The different, quaternary salts obtained on methylation of the corresponding 1,6-bis(dimethylamino) derivatives with methyl iodide (aiming at the structure of epi-allo-muscarine) showed no muscarine-like, biological activity.     

A lipopolysaccharide fromAspergillus flavus conidia

A lipopolysaccharide was isolated by extraction ofAspergillus flavus conidia with 45 % phenol at 68–70 °C. Quantitative analysis revealed 7 % nucleic acids, 5.5 % proteins, 46 % polysaccharides and 49 % lipids, of which 12 % were covalently bound. Glucose, mannose, galactose and fucose were detected as monosaccharide components of the polysaccharide moiety by gas chromatography; palmitic acid, stearic acid, oleic acid, linoleic acid and myristic acid were mainly present in the lipidic fraction. This material differs from the bacterial lipopolysaccharides, both in composition of the polysaccharide moiety and representation of fatty acids in the lipidic fraction.     

Heterogeneity of glutamine synthetase polypeptides in Neurospora crassa

Purified preparations of Neurospora crassa glutamine synthetase contain two nonidentical polypeptides that can be separated by acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and 7 M urea. These polypeptides are synthesized both in vivo and in a heterologous cell-free protein-synthesizing system. The data presented indicate that both polypeptides contain an active site for glutamine synthetase activity and suggest that there is not a precursor-product relationship between them.     

Regulation by aromatic amino acids of the biosynthesis of candicidin by Streptomyces griseus

The biosynthesis by Streptomyces griseus of candicidin, an aromatic polyene macrolide antibiotic, was inhibited by L-tryptophan, L-phenylalanine and, to a lesser degree, by L-tyrosine. A mixture of the three aromatic amino acids inhibited candicidin biosynthesis to a greater extent than did each amino acid separately. L-Tryptophan strongly inhibited the incorporation of the labelled precursors propionate or 4-aminobenzoic acid into candicidin. Incorporation of propionate into candicidin was 50% inhibited by 2.5 mM-tryptophan. Inhibition by tryptophan did not require protein synthesis as the same effect was observed in cells in which protein synthesis was prevented by chloramphenicol. The inhibitory effect of L-tryptophan was partially reversed by exogenous 4-aminobenzoic acid suggesting that this effect is exerted at the level of 4-aminobenzoic acid synthase.     

Control of activity states of heart mitochondrial ATPase. Role of the proton-motive force and Ca2+

The ATPase complex of submitochondrial particles exhibits activity transitions that are controlled by the natural ATPase inhibitor (Gómez-Puyou, A., Tuena de Gómez-Puyou, M. and Ernster, L. (1979) Biochim. Biophys. Acta 547, 252–257). The ATPase of intact heart mitochondria also shows reversible activity transitions; the activation reaction is induced by the establishment of electrochemical gradients, whilst the inactivation reaction is driven by collapse of the gradient. In addition it has been observed that the influx of Ca²⁺ into the mitochondria induces a rapid inactivation of the ATPase; this could be due to the transient collapse of the membrane potential in addition to a favorable effect of Ca²⁺-ATP on the association of the ATPase inhibitor peptide to F₁-ATPase. This action of Ca²⁺ may explain why mitochondria utilize respiratory energy for the transport of Ca²⁺ in preference to phosphorylation. It is concluded that the mitochondrial ATPase inhibitor protein may exert a fundamental regulatory function in the utilization of electrochemical gradients.     

Enzyme polymorphism in a population of Calomys musculinus (Rodentia,Cricetidae)

NAD-linked lactate, malate, glycerophosphate, alcohol and nonspecific dehydrogenases, aspartate aminotransferases, and soluble esterases from extracts of tissues of individuals from a wild population of Calomys musculinus (Rodentia, Cricetidae) have been analyzed by means of starch gel electrophoresis and specific staining. Allelic frequencies and heterozygosity have been determined. Mendelian inheritance of some of the variants detected was confirmed by breeding experiments. Ten out of fifteen (66.6%) of the genetic loci investigated presented polymorphism. Mean heterozygosity per locus was very high (H=0.2014, se 0.046).This work has been supported, in part, by grants from the Secretaria de Ciencia y Tecnología de la Nación (National Program for Endemic Diseases) and from the Fundación Emilio Ocampo. C. N. G. is a Fellow and A. B. a Career Investigator of the Consejo Nacional de Investigaciones Científicas y Técnicas of Argentina.     

Microfluorometric filter determination of DNA in sucrose density gradients

A simple filter method for the fluorometric estimation of DNA in alkaline and neutral sucrose gradient fractions with DABA·2HCl is discussed. Alpha-450 membrane filters of regenerated cellulose (Gelman Instrument Co., Ann Arbor, Michigan) were used. In the proposed method washing of the DNA precipitate as well as the reaction with DABA·2HCl were performed directly on the filters, thus avoiding repeated washing and centrifugations of DNA precipitates applied hitherto in analogous fluorometric techniques. A good coincidence of the results concerning localization of DNA sedimentation profiles determined by radioisotopic and fluorometric methods was obtained. The method is very convenient for DNA estimation in alkaline and neutral sucrose density gradient fractions obtained by ultracentrifugation of DNA of nonproliferating cells where DNA labeling is very difficult, and in the case of human lymphocytes even impossible without stimulation for blastic transformation. Its other advantages are a considerably simplified procedure and a higher precision with respect to other fluorometric methods for determination of DNA.     

Perchlorate binding to cytochrome c: a magnetic and optical study

1. The effects of perchlorate on cytochrome c have been investigated by 1H and 35Cl NMR, electron paramagnetic resonance and optical spectroscopy. 2. The pK values for the formation and disappearance of the major alkaline conformation were found to be displaced from 9.3 to 8.3 and from 10.4 to 10.9, respectively. The displacement was dependent on the ClO4(-) concentration below 0.1 M. 3. Competition experiments between perchlorate and chloride show that ClO4(-) binds both to the neutral and alkaline forms but with a higher affinity for the latter. The appearance of a new binding site in the alkaline form accounts for the markedly enhanced relaxation rate of 35ClO4(-) in this pH range. Complex formation between cyanide and the alkaline species results in the loss of this binding site, which probably is located close to or within the heme crevice. 4. The neutral form of ferricytochrome c also binds perchlorate strongly as evidence by the unique appearance of a high-spin signal dependent on pH and perchlorate concentration. This signal disappears with the same pK value as the neutral form. The effects of perchlorate on cytochrome c are due to specific binding of this ion.