Antimitotics induced cardiolipin cluster formation possible role in mitochondrial enzyme inactivation

We demonstrate here that drugs which inactivate cytochrome c oxidase are able to segregate cardiolipin essential for the enzyme activity, in a separate phase inaccessible for the enzyme. A molecular explanation of the drug-induced aggregation process is proposed.     

Molecular aspects of the bilayer stabilization induced by poly(l-lysines) of varying size in cardiolipin liposomes

The interaction between poly(l-lysines) of varying size with cardiolipin was investigated via binding assays, X-ray diffraction, freeze-fracture electron microscopy, and ³¹P- and ¹³C-NMR. Binding of polylysines to the lipid only occurred when three or more lysine residues were present per molecule. The strength of the binding was highly dependent on the polymerization degree, suggesting a cooperative interaction of the lysines within the polymer. Upon binding, a structural reorganization of the lipids takes place, resulting in a closely packed multilamellar system in which the polylysines are sandwiched in between subsequent bilayers. Acyl chain motion is reduced in these liquid-crystalline peptide-lipid complexes. From competition experiments with Ca²⁺ it could be concluded that when the affinity of the polylysine for cardiolipin was much larger than that of Ca²⁺, a lamellar polylysine-lipid complex was formed, irrespective of whether an excess of Ca²⁺ was added prior to or after the polypeptide. When the affinity of the polylysine for cardiolipin was less or of the same order as that of Ca²⁺, the lipid was organized in the hexagonal H_II phase in the presence of Ca²⁺. These results are discussed in the light of the peptide specificity of bilayer (de)stabilization in cardiolipin model membranes.     

Phospholipid metabolism in Mycobacterium smegmatis ATCC 607 grown at 37° C and 27° C

The rate of synthesis and degradation of phospholipids in Mycobacterium smegmatis ATCC 607, grown at 27° C and 37° C was studied by incorporation of ³²P into phospholipids and chase of radioactivity of the pulse-labelled phospholipids. A relatively low rate of synthesis and degradation of phospholipids in cells growth at 27° C was observed as compared to those grown at 37° C. Phosphatidylethanolamine (PE) had the maximum turnover at 37° C. However, at 27° C, cardiolipin (CL) showed a turnover rate higher than PE. Phosphatidylinositol mannosides (PIMs) were metabolically more active at 37° C than at 27° C. The differences in metabolic activity of the phospholipids at the two temperatures have been discussed.     

Diglyceride kinase from Escherichia coli: Modulation of enzyme activity by glycosphingolipids

Diglyceride kinase was purified from membranes of Escherichia coli K-12 using organic solvents. The enzyme apoprotein depended on lipids, such as cardiolipin (diphosphatidylglycerol), phosphatidylcholine or 1-monooleoylglycerol, for activity with 1,2-dipalmitoylglycerol. Mixed brain cerebrosides and gangliosides as well as defined ganglioside fractions and synthetic lactocerebroside were devoid of lipid cofactor activity. However, all these glycosphingolipids were strong inhibitors of activation by phosphatidylcholine. When cardiolipin was used as lipid activator with the detergent, Triton X-100, as solubilizing agent, the addition of mixed or purified gangliosides first (at about 0.4 mM) resulted in additional activation, but higher ganglioside concentrations were strongly inhibitory. Both effects were absolutely dependent on the presence of lipid-bound sialic acid and were not given by cerebrosides, by free sialic acid or by sialyl-lactose. The stimulating and inhibitory effects of glycosphingolipids could also be demonstrated when 1-monooleoylglycerol was used as substrate, lipid activator and solubilizing agent at the same time. The modulation of kinase activity by glycosphingolipids is discussed at the level of lipid/protein interactions.     

心磷脂-细胞色素C体系CD谱的研究

 线粒体内膜中含有特异的心磷脂是细胞色素C氧化酶活性的必需脂。本工作测定了心磷脂脂质体对细胞色素C溶液园二色(CD)谱的影响,发现心磷脂可引起血色素铁的氧化,并使其轴向配位场强的对称性下降。提示心磷脂可能参与酶和底物之间的电子转移过程。     

Staphylococcus aureus dry stress survivors have a heritable fitness advantage in subsequent dry exposure

Staphylococcus aureus is a major cause of hospital-acquired infections. The ability to survive on abiotic surfaces is an important characteristic that facilitates transmission between human hosts. We found that S. aureus survivors of dry surface incubation are resistant to subsequent dry stress exposure. Survivors also had reduced sensitivity to the disinfectant chlorhexidine gluconate, but not to ethanol. By using a set of mutants in cardiolipin synthase genes, we further demonstrated that the housekeeping cardiolipin synthase, Cls2, was significant for survival on dry surface. Taken together, this study provides insights into S. aureus survival outside of a host.     

Surfactin production enhances the level of cardiolipin in the cytoplasmic membrane of Bacillus subtilis

Surfactin is a cyclic lipopeptide antibiotic that disturbs the integrity of the cytoplasmic membrane. In this study, the role of membrane lipids in the adaptation and possible surfactin tolerance of the surfactin producer Bacillus subtilis ATCC 21332 was investigated. During a 1-day cultivation, the phospholipids of the cell membrane were analyzed at the selected time points, which covered both the early and late stationary phases of growth, when surfactin concentration in the medium gradually rose from 2 to 84 μmol·l^− 1. During this time period, the phospholipid composition of the surfactin producer's membrane (Sf⁺) was compared to that of its non-producing mutant (Sf⁻). Substantial modifications of the polar head group region in response to the presence of surfactin were found, while the fatty acid content remained unaffected. Simultaneously with surfactin production, a progressive accumulation up to 22% of the stress phospholipid cardiolipin was determined in the Sf⁺ membrane, whereas the proportion of phosphatidylethanolamine remained constant. At 24 h, cardiolipin was found to be the second major phospholipid of the membrane. In parallel, the Laurdan generalized polarization reported an increasing rigidity of the lipid bilayer. We concluded that an enhanced level of cardiolipin is responsible for the membrane rigidification that hinders the fluidizing effect of surfactin. At the same time cardiolipin, due to its negative charge, may also prevent the surfactin-membrane interaction or surfactin pore formation activity.     

A cell-based high-throughput screen identifies tyrphostin AG 879 as an inhibitor of animal cell phospholipid and fatty acid biosynthesis

Inhibition of animal cell phospholipid biosynthesis has been proposed for anticancer and antiviral therapies. Using CHOK1 derived cell lines, we have developed and used a cell-based high-throughput procedure to screen a 1280 compound, small molecule library for inhibitors of phospholipid biosynthesis. We identified tyrphostin AG 879 (AG879), which inhibited phospholipid biosynthesis by 85–90% at a concentration of 10 μM, displaying an IC₅₀ of 1–3 μM. The synthesis of all phospholipid head group classes was heavily affected. Fatty acid biosynthesis was also dramatically inhibited (90%). AG879 inhibited phospholipid biosynthesis in all additional cell lines tested, including MDCK, HUH7, Vero, and HeLa cell lines. In CHO cells, AG879 was cytostatic; cells survived for at least four days during exposure and were able to divide following its removal. AG879 is an inhibitor of receptor tyrosine kinases (RTK) and inhibitors of signaling pathways known to be activated by RTK's also inhibited phospholipid biosynthesis. We speculate that inhibition of RTK by AG879 results in an inhibition of fatty acid biosynthesis with a resulting decrease in phospholipid biosynthesis and that AG879's effect on fatty acid synthesis and/or phospholipid biosynthesis may contribute to its known capacity as an effective antiviral/anticancer agent.     

Alteration of mitochondrial function and cell sensitization to death

Stimulation of cell death is a powerful instrument in the organism’s struggle with cancer. Apoptosis represents one mode of cell death. However, in a variety of tumor cells proapoptotic mechanisms are downregulated, or not properly activated, whereas antiapoptotic mechanisms are upregulated. Mitochondria are known as key players in the regulation of apoptotic pathways. Specifically, permeabilization of the mitochondrial outer membrane and subsequent release of proapoptotic proteins from the intermembrane space are viewed as decisive events in the initiation and/or execution of apoptosis. Disruption of mitochondrial functions by anticancer drugs, which induce oxidative stress, inhibit mitochondrial respiration, or uncouple oxidative phosphorylation, can sensitize mitochondria in these cells and facilitate outer membrane permeabilization.