Showing papers by "John L. Harwood published in 1993"
TL;DR: Changes in the acyl lipid composition of maturing embryos of oilseed rape (Brassica napus cv Shiralee) were quantified in plants cultivated in grow.
89 citations
TL;DR: The distribution of labelling within the fatty acids of individual lipid classes was studied and the results showed that the PLs contained significantly more palmitate and significantly lessoleate than the TAGs for embryos at all stages of development.
54 citations
TL;DR: The lipid metabolism in Ascophyllum nodosum was more sensitive to raised Cu2+ levels than in Fucus vesiculosus, and very long chain fatty acids (arachidic, behenic) were increasingly labelled with time.
Abstract: Lipid metabolism and environmental effects on this process have been studied in the marine brown algae Fucus vesiculosus and Ascophyllum nodosum. These algae showed very similar patterns of lipid metabolism during 24 h incubations. Labelling from [1-C-14]acetate showed the major labelled lipids to be the beta-alanine ether lipid and the neutral lipid fraction in both algae. Of the glycolipids, only sulphoquinovosyldiacylglycerol was well labelled and the phosphoglycerides were all poorly labelled. The major labelled fatty acids were palmitate and oleate, again in both algae, although Fucus vesiculosus also showed significant labelling of stearate and behenate. Although the amount of fatty acid labelling increased with time, the proportion of label in palmitate and oleate remained approximately constant. Very long chain fatty acids (arachidic, behenic) were increasingly labelled with time. Lowered incubation temperatures decreased labelling of the saturated fatty acids. Cu2+ increased the proportion of oleate labelled in both algae, and of linoleate in Fucus vesiculosus. This cation decreased the percentage labelling of stearate and myristate in Ascophyllum nodosum. Lipid metabolism in Ascophyllum nodosum was more sensitive to raised Cu2+ levels than in Fucus vesiculosus.
38 citations
TL;DR: The activity of microsomal delta 12-desaturase in Acanthamoeba castellanii was increased after growing cultures were chilled from the optimal growth temperature (30 degrees C) to 15 degrees C.
Abstract: The activity of microsomal delta 12-desaturase in Acanthamoeba castellanii was increased after growing cultures were chilled from the optimal growth temperature (30 degrees C) to 15 degrees C. This increase was detectable in microsomes isolated from organisms subjected to only 10 min chilling. The mechanism of induction was investigated. The increase in activity on chilling was greatly reduced when protein synthesis was blocked before the temperature shift. Thus the major mechanism for the induction of delta 12-desaturase is increased protein synthesis. delta 12-Desaturase activity was higher when assayed at 20 degrees C than when assayed at 30 degrees C, but these changes were not due to the increased solubility of O2 at 20 degrees C. The major substrate of delta 12-desaturase was found to be 1-acyl-2-oleoyl phosphatidylcholine.
31 citations
TL;DR: There appears to be a rapid induction of delta 12-desaturase activity in A. castellanii after a downward shift in growth temperature.
Abstract: A method has been developed for the separation of the major membrane fractions of Acanthamoeba castellanii after growth at different temperatures. The acyl-lipid compositions of individual membrane fractions, microsomal membranes, plasma membrane and mitochondria were analysed after a shift in culture temperature from 30 degrees C to 15 degrees C. The major change in lipid composition observed was an alteration in the relative proportions of oleate and linoleate. This reciprocal change was seen in all the membrane fractions, but occurred most rapidly in the phosphatidylcholine of the microsomal fraction. Thus, there appears to be a rapid induction of delta 12-desaturase activity in A. castellanii after a downward shift in growth temperature. Changes were also seen in the proportions of the n-6 C20 fatty acids, with a decrease in the proportions of icosadienoate and increases of icosatrienoate and arachidonate. However, unlike the alteration in oleate/linoleate ratios, this change was not seen in all the individual lipids of each membrane fraction.
30 citations
TL;DR: Heterotrophic olive callus was characterized by its ability to accumulate triacylglycerol rich in oleate, a situation comparable to developing olive fruit and acyl lipid composition varied according to the state of differentiation and to incubation temperature.
Abstract: Differentiation in olive callus cultures was induced by changing the plant growth regulator content, particularly 2,4-dichlorophenoxyacetic acid, in the growth media at 25 o C. These cultures have been maintained for an extended period with low polyploid nuclei levels. Analysis of olive callus cultures indicated that the acyl lipid composition varied according to the state of differentiation and to incubation temperature. Heterotrophic olive callus was characterized by its ability to accumulate triacylglycerol rich in oleate, a situation comparable to developing olive fruit. In fact, oleate-rich triacylglycerol was enhanced in heterotrophic callus cultured at 35 o C
27 citations
TL;DR: Changes in the relative labelling of triacylglycerols (TAGs) and DAGs in embryos of different ages suggested that diacyl glycerol acyltransferase (DAGAT) could exert significant flux control during periods of rapid lipid synthesis.
27 citations
TL;DR: The steady-state velocity of the enzyme increased in a markedly non-linear fashion with increasing enzyme concentration, indicating that the extent of dissociation of an intermediate in the reaction to free diaminovalerate and the pyridoxaldimine form of thease depends upon the concentration of the enzymes.
Abstract: Glutamate semialdehyde aminotransferase (glutamate-1-semialdehyde 2,1-aminomutase; EC 5.4.3.8) was converted into its pyridoxaldimine form by exhaustive replacement of endogenous pyridoxamine phosphate with pyridoxal phosphate. The isomerization of glutamate 1-semialdehyde to 5-aminolaevulinate by this form of the enzyme followed an accelerating time course which indicated that the enzyme initially had no activity but was converted into the active pyridoxamine phosphate form in an exponential process characterized by a rate constant (k) of 0.027 s-1. The pyridoxaldimine form of the enzyme was converted rapidly into the pyridoxamine form by (S)-4-aminohex-5-enoate and much more slowly by 4-aminobutyrate. The steady-state velocity of the enzyme increased in a markedly non-linear fashion with increasing enzyme concentration, indicating that the extent of dissociation of an intermediate in the reaction to free diaminovalerate and the pyridoxaldimine form of the enzyme depends upon the concentration of the enzyme.
19 citations
TL;DR: Acyl-thioesterase activity has been examined in plastid preparations isolated from two developmental stages in cocoa reflecting low levels of activity at low (105 days post anth) levels.
18 citations
TL;DR: The results suggest that IFN-gamma has a direct effect on the activity of enzymes controlling fatty acid turnover in phospholipids, which altered uptake and turnover of unsaturated fatty acids may have important consequences for the subsequent activation of macrophages by endotoxin.
15 citations
TL;DR: It is demonstrated that there is no relationship between Dialysate phospholipid levels and the adequacy of filtration, although it corroborates previous reports of an inverse correlation between time on CAPD and dialysate lipid concentrations.
Abstract: A qualitative and quantitative study was undertaken to determine the lipid composition of dialysate effluent from patients maintained on continuous ambulatory peritoneal dialysis (CAPD). Effluent, after a 4-h, 2.27% dextrose dwell, was collected on ice, centrifuged and extracted for lipids with chloroform and methanol. Lipids were separated and identified by thin layer chromatography, and the constituent fatty acids were quantitated by gas liquid chromatography. Effluents from 10 patients were assayed at the commencement of CAPD treatment and again after 6 months of therapy. There was a significant fall in phosphatidylcholine and phospholipid concentrations (P
TL;DR: Labelling from [1-C-14]acetate indicated that oleate was the major product of de novo fatty acid synthesis and at higher incubation temperatures more radiolabel was incorporated into saturated fatty acids than at lower temperatures.
Journal Article•
TL;DR: Acyl carrier protein (ACP) is a small protein which is essential for fatty acid synthesis and can be used as a substitute for plant ACP in many assays for plant fatty acid synthetase activity.
Abstract: Acyl carrier protein (ACP) from E. coli was cloned using degenerate PCR. The resulting clone was sequenced and translated to give a product which had the known amino acid sequence of E. coli ACP. The insert obtained was cloned into various expression systems, but no overexpression was obtained and sequencing of the insert revealed'the presence of mutations
TL;DR: E coli ACP can be used as a substitute for plant ACP in many assays for plant fatty acid synthetase activity; indeed it sometimes proves more active in vitro than the native plantACP as mentioned in this paper, which is useful because such large quantities of plant material are required to isolate relatively small amounts of protein.
Abstract: Acyl carrier protein (ACP) is a small (9kDa) protein which is essential for fatty acid synthesis In bacteria [ l ] and plants [2] which use type I1 dissociable fatty acid synthetases, acyl chains are esterified to ACP through the 4'-phosphopantetheine prosthetic group, and ACP acts as a carrier during fatty acid synthesis ACPs from bacteria and plants are very similar All are small acidic proteins with conserved regions particularly around the serine residue through which the prosthetic group is attached The similarity of E coli ACP to many plant ACPs is such that E coli ACP can be used as a substitute for plant ACP in many assays for plant fatty acid synthetase activity; indeed it sometimes proves more active in vitro than the native plant ACP [3] This is useful because such large quantities of plant material are required to isolate relatively small amounts of protein It is therefore easier to isolate E coli ACP E coli contains only one ACP isoform and also has the advantage of being active in many plant system, whereas plant ACPs often do not work in other plant systems, and different and specific isoforms occur in different tissues [4] It is possible to isolate 20-30mg of ACP from 200g of E coli wet cell paste in 3 days However, assaying plant fatty acid synthesis enzymes requires large quantities of ACP and therefore,
Journal Article•
TL;DR: In this article, the authors defined any changes in wheat grain acyl lipids as a result of alterations in environmental growth conditions and found that elevated atmospheric carbon dioxide levels enhanced the accumulation of grain acyl lipids, whereas elevated temperature had the opposite effect.
Abstract: Few studies have been conducted in order to investigate comprehensively the «greenhouse effect» on wheat growth and even less on the alterations to the lipid content of mature wheat grains which are induced by such environmental changes. The objective of this study was to define any changes in wheat grain acyl lipids as a result of alterations in environmental growth conditions. In fact, qualitative and quantitative changes were observed. Most noticeable was the observation that elevated atmospheric carbon dioxide levels enhanced the accumulation of grain acyl lipids, whereas elevated temperature had the opposite effect. Marked alterations in the acyl moieties of the monoacyl phospholipids (starch-located) were also observed
01 Mar 1993
TL;DR: Acyl carrier protein (ACP) from E. coli was cloned using degenerate PCR and the resulting clone was sequenced and translated to give a product which had the known amino acid sequence of E. bacteria ACP.
Abstract: Acyl carrier protein (ACP) from E. coli was cloned using degenerate PCR. The resulting clone was sequenced and translated to give a product which had the known amino acid sequence of E. coli ACP. The insert obtained was cloned into various expression systems, but no overexpression was obtained and sequencing of the insert revealed the presence of mutations.