Phospholipid oxidation catalyzed by cytochrome c in liposomes

Oxidation of liposome phospholipids has been studied in the presence of cytochrome c. Sonicated vesicles of soya bean or egg-yolk lipids, or purified phospholipid preparations, were treated with oxidized cytochrome c at a 10:4 lipid/protein ratio (w/w). Lipid peroxidation was examined by oxygen polarography, gas-liquid chromatography (GLC) and the thiobarbituric acid test. Oxidized, but not reduced, cytochrome effectively catalyzes lipid oxidation under these conditions. Oxygen consumption and disappearance of unsaturated fatty acids follow closely similar patterns, the O2 consumption rate showing a maximum (1.53 mol O2/min per mol heme) shortly before fatty acid loss reaches its peak. GLC and O2 consumption data suggest that monohydroperoxides are the most abundant oxidized species in the system. The thiobarbituric acid reaction, however, appears only to be of qualitative value in peroxidation studies. In order to test the mechanism through which oxidation occurs in our system, the effect of liposome composition and the presence of antioxidants was tested, both on cytochrome c binding to bilayers and on O2 consumption. Oxidized and reduced cytochrome c bind the lipid bilayers with similar affinity, but only the oxidized form is active in autoxidation. Antioxidants do not modify either cytochrome c binding to sonicated liposomes. Lipid composition does influence considerably cytochrome binding, and O2 consumption is correspondingly altered. Studies with various antioxidants and inhibitors suggest that both free radicals and singlet oxygen may be involved in the process under study.     

A note on a modified membrane-Bovis agar for the enumeration of Streptococcus bovis by membrane filtration

The effect of dilution and temperature on the antibacterial properties of potassium sorbate was determined. The time taken to kill a standard inoculum of Escherichia coli was increased considerably after either dilution of the preservative or lowering of the temperature. The value for the concentration exponent, eta, was approximately 3 and that for the temperature coefficient, Q10, was 2.3.     

Evidence for the subcellular localization and specificity of chlordane inhibition in the marine bacterium Aeromonas proteolytica.

Sublethal levels (10 to 100 micrograms/ml) of the chlorinated insecticide chlordane (1,2,4,5,6,7,8,8-octachloro-3a,4,7,7a-tetrahydro-4,7-methanoindan) were introduced into the growth medium of the marine bacterium, Aeromonas proteolytica. Chlordane inhibited the synthesis of an extracellular endopeptidase by almost 40% but exhibited no such inhibition of the extracellular aminopeptidase also produced during the growth cycle. Studied with 14C-labeled chlordane demonstrated that the insecticide was not biologically degraded under the test conditions used and that up to 75% of the recoverable chlordane was cell associated within 48 h. Studied with uniformly labeled L[14C]valine and [2-14C]uracil established that neither the transport nor the incorporation of these protein and ribonucleic acid precursors was inhibited by chlordane. Separation of the membrane fractions using isopycnic centrifugation localized 14C-labeled chlordane in the cytoplasmic membrane. Also, chlordane inhibited the membrane-bound adenosine 5'-triphosphatase while the soluble (released) form of this enzyme remained unaffected. These data indicate that chlordane resides in the cytoplasmic membrane and may cause specific alterations in membrane-associated activities.     

The first seven amino acids encoded by the v-src oncogene act as a myristylation signal: lysine 7 is a critical determinant.

The transforming protein of Rous sarcoma virus, pp60v-src, is covalently coupled to myristic acid by an amide linkage to glycine 2. Myristylation promotes the association of pp60v-src with cellular membranes, and this subcellular location is essential for transforming activity. The findings presented here, in conjunction with the previous reports of others, imply that the seventh amino acid encoded by v-src might be important in the myristylation reaction. Replacement of lysine 7 by asparagine greatly reduced the myristylation, membrane association, and transforming activity of pp60v-src. In contrast, substitution of arginine at residue 7 had no effect on any of these properties of pp60v-src. Addition of amino acids 1 to 7 encoded by v-src was sufficient to cause myristylation of a src-pyruvate kinase fusion protein. We conclude that the recognition sequence for myristylation of pp60v-src comprises amino acids 1 to 7 and that lysine 7 is a critical component of this sequence.