Effect of ammonium ions on spiramycin biosynthesis in Streptomyces ambofaciens

The physiological significance of trans unsaturated fatty acids, which are constituents of membrane lipids of the phenol-degrading bacterium Pseudomonas putita P8, was studied. The addition of phenol or phenol derivatives to the cells induced the formation of trans unsaturated fatty acids, yielding an overall maximal amount of 41.3% of total fatty acids. The inhibition of de-novo lipid synthesis by cerulenin prevented the change in the degree of saturation in the lipids. However, the cells could still respond to phenols with an amplified conversion of cis into trans unsaturated fatty acids, which is apparently a post-synthesis mechanism of isomerization of the double bond. The cis/trans conversion correlated with growth inhibition induced by toxic concentrations of 4-chlorophenol, whereas only growing cells were able to change the degree of saturation. In cells that were protected against phenol by immobilization in calcium alginate, the conversion of cis into trans fatty acids occurred at higher toxin concentrations compared with free cells. Cells entering the stationary growth phase increased the prodortion of saturated to unsaturated fatty acids but maintained a constant trans/cis ratio.P. putida P8 reacted to an increase or decrease in the growth temperature with an appropriate change in the ratio of saturated to unsaturated fatty acids and in cells inhibited by cerulenin with a change in the trans/cis ratio. This study shows that the physiological role of the cis/trans conversion is probably the regulation of membrane fluidity when the most important mechanism for this, the modification of the degree of saturation, cannot by used by the cells due to inhibition of growth and lipid biosynthesis. Correspondence to: H. Keweloh     

Formation of trans Fatty Acids Is Not Involved in Growth-Linked Membrane Adaptation of Pseudomonas putida

Fatty acid compositions in growing and resting cells of several strains of Pseudomonas putida (P8, NCTC 10936, and KT 2440) were studied, with a focus on alterations of the saturation degree, cis-trans isomerization, and cyclopropane formation. The fatty acid compositions of the strains were very similar under comparable growth conditions, but surprisingly, and contrary to earlier reports, trans fatty acids were not found in either exponentially growing cells or stationary-phase cells. During the transition from growth to the starvation state, cyclopropane fatty acids were preferentially formed, an increase in the saturation degree of fatty acids was observed, and larger amounts of hydroxy fatty acids were detected. A lowered saturation degree and concomitant higher membrane fluidity seemed to be optimal for substrate uptake and growth. The incubation of cells under nongrowth conditions rapidly led to the formation of trans fatty acids. We show that harvesting and sample preparation for analysis could provoke the enzyme-catalyzed formation of trans fatty acids. Freeze-thawing of resting cells and increased temperatures accelerated the formation of trans fatty acids. We demonstrate that cis-trans isomerization only occurred in cells that were subjected to an abrupt disturbance without having the possibility of adapting to the changed conditions by the de novo synthesis of fatty acids. The cis-trans isomerization reaction was in competition with the cis-to-cyclopropane fatty acid conversion. The potential for the formation of trans fatty acids depended on the cyclopropane content that was already present.     

Phase transition behaviour of artificial liposomes composed of phosphatidylcholines acylated with cyclopropane fatty acids

Phase transitions of liposomes composed of synthetic phosphatidylcholines acylated with the cyclopropane fatty acids, lactobacillic and dihydrosterculic acid, were studied by differential scanning calorimetry. Transition temperatures were approx. 16°C higher than for phosphatidylcholines acylated with the corresponding unsaturated fatty acids, cis-vaccenic and oleic acid. Though our transition temperatures were all several degrees lower than those determined by Silvius and McElhaney ((1979) Chem. Phys. Lipids 25, 125–134), the increase produced by replacement of the double bond with a cyclopropane ring was the same. We propose that this replacement, through its effect on membrane fluidity, may serve to regulate the activity of membrane-associated processes such as transport.     

Physiological state of microbial communities and mats of the bottom sediments in the marine shallow-water hydrothermal ecosystem of Kraternaya Bight (Kurile Islands)

The physiological state of littoral and sublittoral microbial communities in a marine shallow-water hydrothermal ecosystem (Kraternaya Bight) was studied using lipid biomarkers. The ratio trans/cis (n-7) isomers of monoenic fatty acids (FAs) of polar lipids in intertidal and subtidal algobacterial and bacterial mats of the bight exceeded 0.1 significantly; this indicated a stress state in bacteria. No concomitant increase was found in the ratio of cyclopropane fatty acids to 16: 1 and 18: 1 (n-7) cis monoenic fatty acids. In bottom sediments, the ratio trans/cis (n-7) isomers of monoenic fatty acids was below 0.1. A positive correlation (r = 0.71) was revealed between the ratio trans/cis isomers of (n-7) monoenic fatty acids and the content of saturated fatty acids.     

βVI Turns in peptides and proteins: A model peptide mimicry

To investigate the role of proline in defining β turn conformations within cyclic hexa- and pentapeptides we synthesized and determined the conformations of a series of L - and D -proline-containing peptides by means of 2D NMR spectroscopy and restrained molecular dynamics simulations. Due to cis/trans isomerism the L -proline peptides adopt at least two different conformations that are analyzed and compared to the structures of the corresponding D -proline peptides. The cis conformations of the compounds cyclo(-Pro-Ala-Ala-Pro-Ala-Ala-), cyclo(-Arg-Gly-Asp-Phe-Pro-Gly-), cyclo(-Arg-Gly-Asp-Phe-Pro-Ala-), cyclo(-Pro-Ala-Ala-Ala-Ala--), and cyclo(-Pro-Ala-Pro-Ala-Ala-) form uncommon βVI turns that mimic the turn geometries found in crystallographically refined protein structures at such a detailed level that even preferred side chain orientations are reproduced. The ratios of the cis/trans isomers are analyzed in terms of the steric demand of the proline-following residue. The conformational details derived from this study illustrate the importance of the examination of small model compounds derived from protein loop regions, especially if bioactive recognition sequences, such as RGD (Arg-Gly-Asp), are incorporated. © 1993 Wiley-Liss, Inc.     

Does overwinter temperature affect maternal body composition and egg traits in yellow perch Perca flavescens?

Female yellow perch Perca flavescens exposed to three overwinter temperature regimes (4, 8 and 13° C) for 150 days spawned in markedly different proportions upon spring warming (37% of females in 4° C v. 64 and 91% in 8 and 13° C treatments, respectively), but exhibited no differences in fecundity, egg size or egg lipid content. Females held at 4° C also exhibited less within‐clutch egg size variation than females held at 13° C. Moreover, eggs differed among temperature treatments in the overall proportions of 18 fatty acids, with the colder treatments resulting in potentially higher quality eggs containing more of the unsaturated fatty acids C16:1, C22:6‐n3 and C18:2 cis. Female somatic condition also varied with temperature. Maternal somatic growth and protein content increased while lipid content decreased in 13° C compared to the colder treatments. There were, however, no differences among treatments in the fatty acid composition of maternal muscle. These results suggest that the temperatures experienced during winter may be less influential to P. flavescens egg size or number, which may exhibit relatively little plasticity in this species, but can alter both the number of females that spawn and the overall composition of eggs and maternal somatic tissues, which may have implications for future reproductive success.     

Synthesis of trans unsaturated fatty acids in Pseudomonas putida P8 by direct isomerization of the double bond of lipids

The phospholipids of Pseudomonas putida P8 contain monounsaturated fatty acids in the cis and trans configuration. Cells of this phenol-degrading bacterium change the proportions of these isomers in response to the addition or elimination of a membrane active compound such as 4-chlorophenol. This study undoubtedly reveals that the cis unsaturated fatty acids are directly converted into trans isomers without involvement of de novo synthesis of fatty acids. Oleic acid, which cannot be synthesized by this bacterium, was incorporated as a cis unsaturated fatty acid marker in the membrane lipids of growing cells. The conversion of this fatty acid into the corresponding trans isomer was demonstrated by gas chromatographic-mass spectrometric analysis and use of ¹⁴C-labeled oleic acid. Separation and isolation of the cellular membranes showed that the fatty acid isomerase is located in the cytoplasmic membrane of P. putida P8.Abbreviation 4-CP 4-chlorophenol     

Isolation and optimisation of the oleaginous yeast Sporobolomyces roseus for biosynthesis of 13C isotopically labelled 18-carbon unsaturated fatty acids and trans 18:1 and 18:2 derivatives through synthesis

An oleaginous and psychrotrophic strain (F38-3) of Sporobolomyces roseus Kluyver & van Niel was isolated from a salt marsh environment in Nova Scotia, Canada following a screening program to select for high producers of 18-carbon unsaturated fatty acids. Fatty acid production was characterised as a function of temperature at 20 g glucose L⁻¹, and optimal yields were obtained at 14°C, achieving 5.7 g dw biomass and 39.2% total fatty acids by dry weight, with 18:1, 18:2 and 18:3 all-cis fatty acids accounting for 49.4%, 14.3% and 6.7% of total fatty acids (TFA), respectively—the highest reported for this species. Production of 18:3 was inversely correlated to growth temperature, rising from 2% of TFA at 30°C to 8.9% at 6°C. Cultivation of isolate F38-3 on universally ¹³C (U-¹³C) labelled glucose and subsequent transesterification and isolation of the fatty acid methyl esters (FAMEs) by preparative chromatography yielded pure, highly ¹³C-enriched (>90%) 18:1, 18:2 and 18:3 all-cis FAMEs. The U-¹³C 18:1 FAME was catalytically converted to U-¹³C 18:1 trans-9 and purified to >99.5% purity. The U-¹³C 18:2 was converted by alkaline isomerisation into a 50/50 mixture of 18:2 cis-9, trans-11 and 18:2 trans-10, cis-12 isomers and purified to >95.0% purity. Overall, 10%, by weight, of labelled glucose fed to isolate F38-3 was recovered as fatty acid methyl esters and 7.5% as 18-carbon unsaturated fats, and the final isomerisation reactions resulted in yields of 80% or greater. The ultimate goal of the work is to develop methodologies to produce ¹³C-labelled metabolic tracers as tools to study the metabolism of trans fats.     

Relationship between Stereoisomerism and Biological Activity of Pyrethroids

(+)-trans-Homochrysanthemic acid, when boiled in dilute sulfuric acid, gives (+)-trans-ε-hydroxy-dihydrohomochrysanthemic acid, m.p. 176–7°, together with (+)-δ, δ-dimethyl-γ-isobutenyl-δ-valerolactone. The formation of optically active lactone from (+)-trans-homochrysanthemic acid provides another cogent evidence for the structure of the lactone previously deduced on the racemic compound.

The Arndt-Eistert reaction of the homo-acids give further higher homologues such as (±)-,(+)-trans-β-(3-isobutenyl-2, 2-dimethylcyclopropane-1)-propionic acids and (±)-cis-3-isobutenyl-2, 2 dimethylcyclobutane-1-acetic acid. Both trans-acids, in boiling dilute sulfuric acid, give the same (±)-γ-(1′, 1′, 4′-trimethyl-pent-2′-enyl)-butyrolactone together with the corresponding hydroxy-acids, optically inactive and active, respectively.

Complete resolution of (±)-trans-homochrysanthemic acid and (±)-trans-β-(3-isobutenyl-2, 2-dimethycyclopropane-1)-propionic acid was achieved by means of optically active α-phenylethylamine.     

Metabolic diversity in biohydrogenation of polyunsaturated fatty acids by lactic acid bacteria involving conjugated fatty acid production

Lactobacillus plantarum AKU 1009a effectively transforms linoleic acid to conjugated linoleic acids of cis-9,trans-11-octadecadienoic acid (18:2) and trans-9,trans-11–18:2. The transformation of various polyunsaturated fatty acids by washed cells of L. plantarum AKU 1009a was investigated. Besides linoleic acid, α-linolenic acid [cis-9,cis-12,cis-15-octadecatrienoic acid (18:3)], γ-linolenic acid (cis-6,cis-9,cis-12–18:3), columbinic acid (trans-5,cis-9,cis-12–18:3), and stearidonic acid [cis-6,cis-9,cis-12,cis-15-octadecatetraenoic acid (18:4)] were found to be transformed. The fatty acids transformed by the strain had the common structure of a C18 fatty acid with the cis-9,cis-12 diene system. Three major fatty acids were produced from α-linolenic acid, which were identified as cis-9,trans-11,cis-15–18:3, trans-9,trans-11,cis-15–18:3, and trans-10,cis-15–18:2. Four major fatty acids were produced from γ-linolenic acid, which were identified as cis-6,cis-9,trans-11–18:3, cis-6,trans-9,trans-11–18:3, cis-6,trans-10–18:2, and trans-10-octadecenoic acid. The strain transformed the cis-9,cis-12 diene system of C18 fatty acids into conjugated diene systems of cis-9,trans-11 and trans-9,trans-11. These conjugated dienes were further saturated into the trans-10 monoene system by the strain. The results provide valuable information for understanding the pathway of biohydrogenation by anaerobic bacteria and for establishing microbial processes for the practical production of conjugated fatty acids, especially those produced from α-linolenic acid and γ-linolenic acid. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Studies of Chrysanthemic Acid

(±) -cis and trans-2,2-dimethyl-3- (2′-cyano-l′-propenyl) cyclopropane carboxylic acids and their optically active forms were synthesized starting from chrysanthemic acids via oximes of 2,2-dimethyl3- (2′-formyl-l′-propenyl) cyclopropane carboxylates. Their allethrolone esters were also prepared which showed insecticidal activity.     

Effects of Feeding Pigs Increasing Levels of C 18:1 Trans Fatty Acids on Fatty Acid Composition of Backfat and Intramuscular Fat as well as Backfat Firmness

Forty Large White pigs were fed from 30kg to 103kg body mass on diets supplemented with 6% of pure high-oleic sunflower oil (HO) or HO plus increasing amounts of partially hydrogenated rape seed oil (HR; 1.85%, 3.70%, 5.55%), containing high levels of j 6 to j 11 C 18:1 trans fatty acid isomers. Increasing dietary C 18: trans fatty acids resulted in a linear increase in C 18:1 trans fatty acids and conjugated linoleic acid (cis-9, trans-11 CLA) in backfat (BF) as well as in neutral lipids (NL) and phospholipids (PL) of M. long. dorsi. Thus, the rate of bioconversion of trans vaccenic acid (TVA) into CLA and incorporation of C 18:1 trans and CLA into pig adipose tissue was not limited up to 25g total C 18:1 trans fatty acids including 3.3g of TVA perkg feed. BF was higher in C 18:1 trans fatty acids and CLA than M. long. dorsi NL and PL. In BF and NL the sum of saturated fatty acids (SFA) increased with increasing dietary amounts of HR, while in PL SFA were reduced. Thus, according to their physical properties, C 18:1 trans fatty acids partly replaced SFA in PL. Firmness of backfat was also significantly increased (P<0.05) with increasing amounts of HR in feed.     

Reorganization of membrane lipids during fast and slow cold hardening in Drosophila melanogaster

Abstract Rapid cold hardening is a naturally occurring phenomenon in insects that is thought to be responsible for increased cold tolerance during diurnal variations in temperature. The underlying physiological mechanisms are still not fully resolved but, in Drosophila melanogaster (Meigen 1830), rapid cold hardening is accompanied by specific changes in the membrane lipid composition. To further understand the link between rapid cold hardening and adjustments in the membrane lipid composition, the present study investigates how different rates of cooling affect thermotolerance and the composition of phospholipid fatty acids. Female Drosophila are cooled gradually from 25 to 0 °C at 0.01, 0.05, 0.1 or 0.5 °C min^?1, respectively, and, subsequently, phospholipid fatty acid composition and survival after a 1‐h cold shock at ?5 °C is measured. The rapid cold hardening treatments all influence cold tolerance differently so that short and intermediate rapid cold hardening treatments (0.05, 0.1 or 0.5 °C min^?1 cooling rates) increase cold shock survival, whereas the slow cooling treatment (0.01 °C min^?1) decreases survival relative to an untreated control. The intermediate rapid cold hardening treatments (0.05 or 0.1 °C min^?1) induce a similar type of response characterized by an increase in the molar percentage of linoleic acid, 18:2(n‐6), at the expense of 16:0 and 18:1(n‐9), which leads to an increase in the degree of unsaturation. The slowest cooling treatment (0.01 °C min^?1) results in a large increase in cis‐16:1(n‐7) and significant reductions in the saturated phospholipid fatty acids 16:0, 18:0 and the unsaturated 16:1(n‐9) and 18:2(n‐6) fatty acids. These changes cause a slight decrease in the average length of the phospholipid fatty acids and an increase in the overall ratio of unsaturated vs. saturated fatty acids. These findings demonstrate that the rate of cooling is important for both the reorganization of membrane lipids, and for the degree of acquired cold tolerance during rapid cold hardening, and they suggest an important role for rapid cold hardening during diurnal rather than seasonal temperature changes.     

Effect of different types of fibre supplemented with sunflower oil on ruminal fermentation and production of conjugated linoleic acids in vitro

Abstract

An in vitro study was conducted to determine the effect of different types of fibre supplemented with sunflower oil on ruminal fermentation and formation of conjugated linoleic acids (CLA) by mixed ruminal microorganisms. Cell wall components extracted from wheat straw (representing lignified fibre), soybean hulls (representing easily digestible fibre), and purified cellulose were used as substrates. Sunflower oil was supplemented at the same level for all three types of fibre. After 24 h of incubation, ruminal fermentation parameters (including 24 h gas production, pH value, concentration of ammonia nitrogen and volatile fatty acids) and the concentration of long chain fatty acids in the culture fluid were determined. Results showed that the type of fibre influenced ruminal fermentation traits and the biohydrogenation of unsaturated C18 fatty acids in vitro. Composition of LCFA and profile of CLA were altered by the fibre type. Compared to the digestible fibre and purified cellulose, lignified fibre significantly increased the production of cis-9, trans-11 CLA and total CLA (sum of cis-9, trans-11 CLA, trans-10, cis-12 CLA, trans-9, trans-11 CLA, and cis-9, cis-11 CLA) by ruminal microorganisms. It was concluded that ruminal fermentation and production of CLA can be affected by the type of dietary fibre.     

Relation between Cyclopropane Fatty Acid Formation and Saturation of Phospholipid Species in Escherichia coli K-12

Alterations in the degrees of saturation of phosphatidylethanolamine, phosphatidylglycerol and cardiolipin of Escherichia coli K–12 were determined after raising or lowering the growth temperature during the exponential growth phase. After raising the growth temperature from 17 to 42°C, the cells continued to grow with increasing degrees of saturation of the three phospholipids. cis-9,10-Methylenehexadecanoic acid increased only in phosphatidylethanolamine. During growth after lowering the growth temperature from 42 to 17°C, no increase was found in cyclopropane fatty acid content of phosphatidylethanolamine, in which cis-vaccenic acid increased. Significance of cyclopropane fatty acid formation in phospholipids was discussed.     

Metabolic Implications of Dietary Trans‐fatty Acids

Dietary trans‐fatty acids are associated with increased risk of cardiovascular disease and have been implicated in the incidence of obesity and type 2 diabetes mellitus (T2DM). It is established that high‐fat saturated diets, relative to low‐fat diets, induce adiposity and whole‐body insulin resistance. Here, we test the hypothesis that markers of an obese, prediabetic state (fatty liver, visceral fat accumulation, insulin resistance) are also worsened with provision of a low‐fat diet containing elaidic acid (18:1t), the predominant trans‐fatty acid isomer found in the human food supply. Male 8‐week‐old Sprague–Dawley rats were fed a 10% trans‐fatty acid enriched (LF‐trans) diet for 8 weeks. At baseline, 3 and 6 weeks, in vivo magnetic resonance spectroscopy (¹H‐MR) assessed intramyocellular lipid (IMCL) and intrahepatic lipid (IHL) content. Euglycemic–hyperinsulinemic clamps (week 8) determined whole‐body and tissue‐specific insulin sensitivity followed by high‐resolution ex vivo ¹H‐NMR to assess tissue biochemistry. Rats fed the LF‐trans diet were in positive energy balance, largely explained by increased energy intake, and showed significantly increased visceral fat and liver lipid accumulation relative to the low‐fat control diet. Net glycogen synthesis was also increased in the LF‐trans group. A reduction in glucose disposal, independent of IMCL accumulation was observed in rats fed the LF‐trans diet, whereas in rats fed a 45% saturated fat (HF‐sat) diet, impaired glucose disposal corresponded to increased IMCL_TA. Neither diet induced an increase in IMCL_soleus. These findings imply that trans‐fatty acids may alter nutrient handling in liver, adipose tissue, and skeletal muscle and that the mechanism by which trans‐fatty acids induce insulin resistance differs from diets enriched with saturated fats.     

The partition of cis-parinaric acid and trans-parinaric acid among aqueous,fluid lipid,and solid lipid phases

Summary In this article, I review the current information concerning the partition of the fluorescent probes, cis-parinaric acid (9, 11, 13, 15-cis, trans, trans, cis-octadecatetraenoic acid) and trans-parinaric acid (9, 11, 13, 15-all trans-octadecatetraenoic acid) among aqueous, solid lipid, and fluid lipid phases. The association of these probes with lipid is described by a mole fraction partition coefficient whose value is typically in the range of 1–5 × 10⁶, a reasonable value in light of partition coefficients for other fatty acids between hydrophobic phases and water. The partition coefficient, in the absence of lipid phase changes, is relatively independent of temperature and only slightly dependent on the total aqueous probe concentration.In lipid samples which contain coexisting fluid and solid phases, trans-parinaric acid preferentially partitions into the solid phase, while cis-parinaric acid distributes nearly equally between fluid and solid phases. This partition behavior probably arises from the molecular shape of the cis and trans parinaric acid isomers. From measurements of the polarization of fluorescence of cis and trans parinaric acid in mixed lipid systems or membranes it is possible to evaluate the proportion of lipid components involved in phase changes or phase separation. From fluorescence energy transfer between protein typtophan residues and the parinaric acid isomers it is possible to gain information about the organization of lipids and proteins in membranes and model systems. I close the review by considering some of the membrane research areas where these probes and their various lipid derivatives may be particularly useful.     

How do bacteria sense and respond to low temperature?

Rigidification of the membrane appears to be the primary signal perceived by a bacterium when exposed to low temperature. The perception and transduction of the signal then occurs through a two-component signal transduction pathway consisting of a membrane-associated sensor and a cytoplasmic response regulator and as a consequence a set of cold-regulated genes are activated. In addition, changes in DNA topology due to change in temperature may also trigger cold-responsive mechanisms. Inducible proteins thus accumulated repair the damage caused by cold stress. For example, the fluidity of the rigidified membrane is restored by altering the levels of saturated and unsaturated fatty acids, by altering the fatty acid chain length, by changing the proportion of cis to trans fatty acids and by changing the proportion of anteiso to iso fatty acids. Bacteria could also achieve membrane fluidity changes by altering the protein content of the membrane and by altering the levels of the type of carotenoids synthesized. Changes in RNA secondary structure, changes in translation and alteration in protein conformation could also act as temperature sensors. This review highlights the various strategies by which bacteria senses low temperature signal and as to how it responds to the change.     

Solvent and side-chain contributions to the two-cis ⇄ all-trans equilibria of cyclic hexapeptides,cyclo(Xxx-Pro-D-Yyy)2

Cyclic hexapeptides of the type cyclo(L -Xxx-L -Pro-D -Yyy)₂ or cyclo(L -Xxx-L -Pro-Gly)₂ exist in solution predominantly in two forms of C₂ average symmetry, one with all-trans peptide bonds and generally well-established conformation, and another with both Xxx-Pro peptide bonds cis. We have been measuring the thermodynamic parameters of this equilibrium using carbon and proton nmr spectroscopy. Data have been obtained for peptides in which Yyy = Gly, D -Ala, or D -Phe, and Xxx = Gly, L -Ala, L -Leu, and L -Val. In a given solvent, stability of the all-trans form decreases (ΔG⁰ increases) as Xxx is changed through the series Gly, L -Ala-, L -Leu, and L -Val, consistent with expected increasing repulsion between the Xxx side chain and the proline δ methylene across the trnas Xxx-Pro bond. Also, for a given set of side chains, the stability of the all-trnas form increases as the polarity of the solvent decreases, consistent with models in which all C?O and N? H groups are accessible for solvation in the two-cis form, but two C?O and two N? H groups are somewhat sequestered in the all-trans form. With the available data it is not possible to identify pure intramolecular (solvent-independent) or pure peptide-bond solvation (side chain-independent) terms in ΔH° or ΔS°, although trends are discernible.