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L O Ingram

Publications and source records attributed to L O Ingram.

At least 109 records · Page 6Linked to original sources
PubMed1983

Lipid composition of Zymomonas mobilis: effects of ethanol and glucose.

Zymomonas mobilis is an alcohol-tolerant microorganism which is potentially useful for the commercial production of ethanol. This organism was found to contain cardiolipin, phosphatidylethanolamine, phosphatidylglycerol, and phosphatidylcholine as major phospholipids. Vaccenic acid was the most abundant fatty acid, with lesser amounts of myristic, palmitic, and palmitoleic acids. No branched-chain or cyclopropane fatty acids were found. Previous studies in our laboratory have shown that ethanol induces the synthesis of phospholipids enriched in vaccenic acid in Escherichia coli (L. O. Ingram, J. Bacteriol. 125:670-678, 1976). The fatty acid composition of Z. mobilis, an obligately ethanol-producing microorganism, represents an extreme of the trend observed in E. coli. In Z. mobilis, vaccenic acid represents over 75% of the acyl chains in the polar membrane lipids. Glucose and ethanol had no major effect on the fatty acid composition of Z. mobilis. However, both glucose and ethanol caused a decrease in phosphatidylethanolamine and phosphatidylglycerol and an increase in cardiolipin and phosphatidylcholine. Ethanol also caused a dose-dependent reduction in the lipid-to-protein ratios of crude membranes. The lipid composition of Z. mobilis may represent an evolutionary adaptation for survival in the presence of ethanol.

Bacteria↗
PubMed1982

Unsaturated fatty acid requirement in Escherichia coli: mechanism of palmitate-induced inhibition of growth of strain WN1.

The minimum requirement for unsaturated fatty acids was investigated in E. coli using a mutant impaired in the synthesis of vaccenic acid. Exogenously supplied palmitic acid was incorporated by this mutant which led to a reduction in the proportion of cellular unsaturated fatty acids. Growth was impaired as the level of saturated fatty acids approached 76% at 37 degree C and 60% at 30 degree C. The basis of this growth inhibition was investigated. Most transport systems and enzymes examined remained active in palmitate-grown cells although the specific activities of glutamate uptake and succinic dehydrogenase were depressed 50%. Fluorescent probes of membrane organization indicated that fluidity decreased with palmitate incorporation. Temperature scans with parinaric acid indicated that rigid lipid domains exist in palmitate-grown cells at their respective growth temperature. Freeze-fracture electron microscopy confirmed the presence of phase separations (particle-free areas) in palmitate-grown cells held at their growth temperature prior to quenching. The extent of this separation into particle-free and particle-enriched domains was equivalent to that induced by a shift to 0 degree C in control cells. The incorporation of palmitate increased nucleotide leakage over threefold. The cytoplasmic enzyme beta-galactosidase was released into the surrounding medium as the concentration of unsaturated fatty acid approached the minimum for a particular growth temperature. Lysis was observed as a decrease in turbidity when cells which had been grown with palmitate were shifted a lower growth temperature. From these results we propose that leakage and partial lysis are the major factors contributing to the apparent decrease in growth rate caused by the excessive incorporation of palmitate. Further, we propose that membrane integrity may determine the minimum requirement for unsaturated fatty acids in E. coli rather than a specific effect on membrane transport and/or membrane-bound enzymes.

Biological Transport, Active↗
PubMed1982

Acute effects of ethanol on biosynthesis and glycosylation of IgGl(kappa) antibody molecules in cultured P3/X63-Ag8 myeloma cells.

Protein synthesis in cultured P3/X63-Ag8 mouse myeloma cells was inhibited by acute exposure to ethanol. However, the synthesis of IgGl antibody, as a percentage of total protein synthesis, increased slightly. Experiments using actinomycin D suggest that the overall inhibition of protein synthesis by ethanol occurs at the translational level. Following an L-[35S]methionine pulse, cultured P3/X63-Ag8 cells contained one light antibody polypeptide and two heavy antibody polypeptides. One of these heavy chains was shown to be the unglycosylated precursor of the other, mature molecule. Only the glycosylated polypeptide is a normal constituent of secreted IgGl antibody. The glycosylation of the immature heavy chains occurred more rapidly during a 1-hr isotopic pulse in cells exposed to ethanol (0.1 v/v % and above), than in unexposed control cells. The observed effects of ethanol on antibody glycosylation may be related to the increased susceptibility of alcoholic patients to infections. Ethanol may also affect the synthesis of other glycoproteins in myeloma cells and other tissues.

Animals↗
PubMed1982

Regulation of fatty acid composition in Escherichia coli: a proposed common mechanism for changes induced by ethanol, chaotropic agents, and a reduction of growth temperature.

Growth of Escherichia coli in the presence of ethanol and chaotropic salts resulted in the synthesis of lipids containing elevated levels of unsaturated fatty acids analogous to the effect of a reduction in growth temperature. Both ethanol and chaotropic agents acted at the level of fatty acid biosynthesis and altered lipid composition by decreasing the proportion of saturated acyl chains available for the synthesis of phospholipids. A reduction in temperature causes similar effects on fatty acid biosynthesis in vivo and in vitro. Ethanol, chaotropic salts, and a decrease in temperature all weaken hydrophobic interactions. Antichaotropic salts antagonized and effects of these treatments on fatty acid synthesis in vitro. These results are consistent with a common mechanism for the effects of chaotropic agents, temperature, and ethanol on fatty acid synthesis. The biosynthesis of saturated and unsaturated acyl chains may be regulated by the strength of hydrophobic interactions. Changes in the strength of hydrophobic interactions could alter enzyme structure, substrate structure, or the equilibrium between the soluble enzymes of fatty acid synthesis and their respective acyl carrier protein substrates.

Escherichia coli↗
PubMed1982

Effects of fatty acid composition on the sensitivity of membrane functions to ethanol in Escherichia coli.

The effects of membrane composition on the sensitivity of membrane functions to ethanol in Escherichia coli have been investigated. The addition of ethanol (0.67M) in vitro did not cause appreciable inhibition of NADH oxidase, D-lactate oxidase or ATPase but caused an 11% to 30% inhibition of succinic dehydrogenase, glutamate uptake, proline uptake and 1ac permease. Although the sensitivities of some of these membrane functions to ethanol varied with membrane composition, none correlated with the changes in sensitivity to killing by ethanol. In contrast, leucine transport was resistant to ethanol (0.67M) in control cells and in cells enriched in vaccenic acid, but was inhibited by 25% in cells grown in palmitic acid. The release of nucleotides was examined as a comparative measure of cellular permeability. Ethanol increased nucleotide leakage. Leakage was reduced in cells grown in vaccenic acid and enhanced in cells enriched in palmitic acid. The addition of MgSO4 (10mM) reduced nucleotide leakage and enhanced survival. Based upon these results, metabolite leakage was proposed as the primary event associated with bacterial inactivation in buffered solutions of ethanol. The increase in acyl chain length is proposed as the beneficial aspect of vaccenic acid incorporation rather than the increase in membrane unsaturation.

Amino Acids↗
PubMed1982

On the relationship between alcohol narcosis and membrane fluidity.

We have examined the relationship between membrane fluidity and the alcohol-induced loss of righting reflex at different temperatures using the fish Gambusia affinis. The potency of ethanol and hexanol increased dramatically with temperature. Ethanol-induced narcosis could be antagonized by a reduction in incubation temperature. Both an increase in temperature and the addition of ethanol caused an increase in membrane fluidity. However, membrane fluidity itself did not correlate with narcosis and was primarily determined by incubation temperature. Narcotic concentrations of ethanol caused a change in fluidity equivalent to less than that caused by a 2 degree increase in temperature while an 8 degree increase in temperature did not induce narcosis. From these studies, we conclude that the ethanol-induced increase in bulk membrane fluidity as measured by diphenyl-hexatriene is not the causal event for narcosis although the magnitude of this change does correlate with the alcohol sensitivity. We have also examined the effects of temperature adaptation on the sensitivity of these animals to ethanol. Summer animals contained higher levels of saturated fatty acids, exhibited a higher temperature range than winter animals and were more resistant to ethanol, providing further evidence for membrane structure as a determinant of alcohol sensitivity.

Animals↗
PubMed1981

Effects of chaotropic and antichaotropic agents on elution of poliovirus adsorbed on membrane filters.

The association of poliovirus with membrane filters results from both electrostatic and hydrophobic interactions. At low pH, electrostatic interactions appear to dominate. However, at high pH, hydrophobic interactions appear to dominate with both Millipore and Zeta plus filters. With both filters, viral elution was prevented at high pH by the presence of antichaotropic salts, which strengthen hydrophobic associations. This effect was antagonized by detergents and by chaotropic salts, which weaken hydrophobic associations. Excellent correlation was observed between viral elution, the solubilization of adenine, and the micelle-formation process in the presence of chaotropic and antichaotropic agents. Such simple measurements of the relative ability of solvents to accommodate hydrophobic groups may be of predictive value in designing methods for concentration and purification of viruses. The hypothesis that hydrophobic interactions are the primary force involved in the stabilization of virus attachment at high pH is consistent with observations that cannot be explained by consideration of electrostatic interactions as the major stabilizing factor.

Adsorption↗
PubMed1981

Lipid synthesis during the Escherichia coli cell cycle.

Lipid synthesis was examined in Escherichia coli cells at different stage of cell division. Exponentially growing cells were pulse-labeled with appropriate isotopes for 0.1 generation time, inactivated, and separated by size on a sucrose gradient. An abrupt increase in the rate of lipid synthesis occurred which was coincident with the initiation of cross walls. In contrast, the rate of protein synthesis during this same interval remained constant, resulting in an increased lipid/protein ratio in dividing cells. No changes in the composition of phospholipid head groups, fatty acids, or phospholipid molecular species were observed in cells at different stages of division. The observed increase in the rate of lipid synthesis may reflect a means by which the activities of membrane-associated enzymes are modulated during cross wall formation.

Bacterial Proteins↗
PubMed1981

Mechanism of lysis of Escherichia coli by ethanol and other chaotropic agents.

Ethanol has been shown to inhibit the assembly of cross-linked peptidoglycan and to induce cell lysis in Escherichia coli. These effects of ethanol appear to result from the weakening of hydrophobic interactions by ethanol rather than from the intercalation of ethanol into membranes. Other chaotropic agents also inhibited cross-linking and induced lysis. The potency of chaotropic anions with regard to this effect followed the expected chaotropic series. Antichaotropic agents, which strengthened hydrophobic interactions, antagonized ethanol-induced lysis. The weakening of hydrophobic interactions by ethanol is proposed as a general mechanism by which ethanol and other chaotropic agents could affect membrane-associated enzyme activities.

Bacteriolysis↗
PubMed1981

Failure of Escherichia coli to alter its fatty acid composition in response to cholesterol-induced changes in membrane fluidity.

Substantial amounts of exogenously supplied cholesterol were incorporated into the membranes of Escherichia coli during growth and caused a large decrease in membrane fluidity. Although no compensatory changes in fatty acid composition were observed, the incorporation of cholesterol did not affect the rate of growth of E. coli or interfere with the changes in fatty acid composition which normally occur during growth at different temperatures.

Cholesterol↗
PubMed1980

Reversible effects of ethanol on E. coli.

Chronic exposure of E. coli to ethanol during growth resulted in major changes in lipid composition. These ethanol-induced changes, a decrease in the proportion of saturated fatty acids, are similar to those which occur following a shift to lower temperature. Products of ethanol metabolism such as acetaldehyde and acetate caused the opposite changes in fatty acid composition. In vivo studies using mutants blocked in lipid synthesis indicated that saturated fatty acid synthesis was the primary target leading to changes in bulk lipid fatty acid composition. This was confirmed in vitro and condensing enzyme II was identified as the probable site of ethanol inhibition. The acute affects of ethanol on the function of two membrane-bound enzymes, Mg++ATPase and lac permease were also examined. In both cases, cells grown in the presence of ethanol. In time-course studies, permease function was restored concurrently with changes in lipid composition. Mutants were isolated which were able to grow in the presence of high levels of ethanol. These mutants displayed exaggerated changes in lipid composition providing evidence that alcohol-resistance and fatty acid changes are related.

Adenosine Triphosphatases↗
PubMed1980

Alcohol tolerance in Escherichia coli.

During growth with ethanol, the proportion of 18:1 fatty acid in the lipids of E. coli increases at the expense of saturated fatty acids. The significance of these changes was investigated in terms of growth and survival in the presence of ethanol. Two approaches were used: (1) A comparison of alcohol tolerance among strains of E. coli with different fatty acid compositions; (2) A comparison of alcohol tolerance using a lipid mutant in which the fatty acid composition was controlled by exogenous supplements. An increase in unsaturated fatty acid content was beneficial for both growth and survival. We conclude that the alcohol-induced changes in the fatty acid composition of E. coli are part of an adaptive response, compensating for some of the harmful effects of this drug.

Cell Membrane↗
PubMed1980

Alcohol-induced changes in the phospholipid molecular species of Escherichia coli.

In Escherichia coli, the additon of ethanol resulted in the synthesis of an increased proportion of phospholipids containing two unsaturated fatty acids. The addition of hexanol resulted in the opposite effect, an increase in the proportion of monounsaturated molecular species. The alcohol-induced changes were quantitatively similar to those caused by changing growth temperature. These results suggest that both adaptation to temperature and alcohol-induced changes in lipid composition share some common regulatory features.

Diglycerides↗
PubMed1980

Differential effects of ethanol and hexanol on the Escherichia coli cell envelope.

Both ethanol and hexanol inhibited the growth of Escherichia coli, but their effects on the organization and composition of the cell envelope were quite different. Hexanol (7.8 x 10(-3) mM) increased membrane fluidity, whereas ethanol (0.67 M) had little effect. During growth in the presence of ethanol, the proportion of unsaturated fatty acids increased. The opposite change was induced by hexanol. Unlike hexanol, growth in the presence of ethanol resulted in the production of un-cross-linked peptidoglycan with subsequent lysis. Salt (0.3 M) protected cells against ethanol-induced lysis but potentiated growth inhibition by hexanol. Mutants isolated for resistance to ethanol-induced lysis synthesized cross-linked peptidoglycan during growth in the presence of ethanol but remained sensitive to hexanol. A general hypothesis was presented to explain the differential effects of ethanol and hexanol. All alcohols are viewed as similar in having both an apolar chain capable of interacting with hydrophobic environments and a hydroxyl function capable of hydrogen bonding. The differential effects of short-chain alcohols may represent effects due to the high molar concentrations of hydrogen bonding groups with an apolar end within the environment. These may replace bound water in some cases. With longer-chain alcohols such as hexanol, the effects of the acyl chain would dominate, and limitations of solubility and cellular integrity would mask these hydroxyl effects.

Bacteriolysis↗
PubMed1978

Inhibition of unsaturated fatty acid synthesis in escherichia coli by the antibiotic cerulenin.

Low concentrations of cerulenin inhibit the growth of Escherichia coli by selectively blocking unsaturated fatty acid synthesis. This inhibition was relieved by unsaturated fatty acid supplements alone but not by saturated fatty acid supplements. The utilization of exogenous unsaturated fatty acids to sustain growth in the presence of cerulenin was confirmed by the analysis of bulk lipid composition. The effects of cerulenin on fatty acid synthesis were examined in vivo by pulse labeling with [14C]acetate and in vitro using [14C]malonyl-coenzyme A. In both cases, unsaturated fatty acid synthesis was inhibited by low concentrations of cerulenin with a stimulation of saturated fatty acid synthesis. Using mutant strains deficient in fatty acid synthesis, the effects of cerulenin on beta-ketoacyl-[acyl-carrier-protein] synthetases I and II were examined. Our results indicate that beta-ketoacyl-[acyl-carrier-protein] synthetase I is more sensitive to inhibition by cerulenin than beta-ketoacyl-[acyl-carrier-protein] synthetase II.

3-Oxoacyl-(Acyl-Carrier-Protein) Synthase↗
PubMed1978

Mechanism of ethanol-induced changes in lipid composition of Escherichia coli: inhibition of saturated fatty acid synthesis in vivo.

The in vivo effects of ethanol on lipid synthesis in Escherichia coli have been examined. Under conditions which uncoupled fatty acid synthesis from phospholipid synthesis, ethanol decreased the amount of saturated fatty acids synthesized but had little effect on the selectivity of their incorporation into phospholipids. In the absence of fatty acid degradation and unsaturated fatty acid synthesis, E. coli was still able to adapt its membrane lipids to ethanol, while the inhibition of total fatty acid synthesis eliminated this response. During growth in the presence of ethanol, strain K1060 (an unsaturated fatty acid auxotroph) incorporated an increased amount of exogenous heptadecanoic acid (17:0) to compensate for the reduction in palmitic acid (16:0) available from biosynthesis. Thus, our results indicate that the reduced levels of saturated fatty acids observed in the phospholipids of E. coli following growth in the presence of ethanol result primarily from a decrease in the amounts of saturated fatty acids available for phospholipid synthesis.

Escherichia coli↗