John L. Harwood

Bio: John L. Harwood is an academic researcher from Cardiff University. The author has contributed to research in topics: Lipid metabolism & Fatty acid. The author has an hindex of 60, co-authored 420 publications receiving 16081 citations. Previous affiliations of John L. Harwood include John L. Scott & Spanish National Research Council.

Informed metabolic engineering of oil crops using control analysis

Abstract: Oil crops are a very important agricultural commodity. Demand for such oils is rising steadily (at more than 5% per year over the last half century). Although the majority of plant oils are used for food or animal feed, there is increasing interest in their use as renewable chemicals for industry. Because of the demonstrated demand for oils and finite agricultural land, attention is focussing on improving productivity. Genetic manipulation of crop plants needs a knowledge of the biosynthetic pathways concerned and how they are regulated. Although there are different ways to acquire much information, metabolic flux and metabolic control analyses are ways to provide quantitative assessments. In this review we describe our experiments using metabolic control analysis on important crops – oil palm, oilseed rape, olive and soybean. Such research provides information for future informed genetic manipulations and we give a successful example of this in oilseed rape (Brassica napus L.).

Oxygen-dependent low-temperature δ12 (n6)-desaturase induction and alteration of fatty acid composition in Acanthamoeba castellanii

Abstract: The influence of dissolved oxygen on the synthesis and activity of δ12-desaturase in Acanthamoeba castellanii was investigated. A decline in oxygen concentration during batch growth at 30° was correlated with a decline in the degree of cellular fatty acid unsaturation. Chilling of early-stationary-phase cultures to 15° led to increased dissolved oxygen levels (from 160 μM), gave no change in cellular fatty acid unsaturation. Measurement of [1-14C]acetate incorporation by oxygen-limited A. castellanii revealed that labelling of the δ12-desaturase product, linoleate (18:2), increased with oxygen concentration. Microsomal levels of the δ12-desaturase enzyme were found to increase by up to 10-fold during aeration of A. castellanii cultures; a transient elevation in oxygen was sufficient to induce δ12-desaturase synthesis that was still fully detectable 1 h later. In addition, the activity of pre-existing δ12-desaturase, measured in isolated microsomal membranes, increased by up to fivefold with increases in the oxygen concentration of assay mixtures. These results demonstrate for the first time that (i) oxygen availability alone can regulate de novo δ12-desaturase synthesis in A. castellanii, and that (ii) oxygen can limit the activity of pre-existing δ12-desaturase. These responses can occur independently of temperature changes.

Re-evaluation of plant sulpholipid labelling from UDP-[14C]glucose in pea chloroplasts.

Abstract: The sulphoquinovosyldiacylglycerol (sulpholipid) zone obtained by TLC of pea chloroplast lipids yields, on hydrolysis, not only sulphoquinovose but also galactose and glucose. Following incorporation from UDP-[(14)C]glucose, the percentages of the total radioactivity in these three sugars were typically 1, 5 and 85%, respectively. The occurrence of the glucolipid causes difficulties in elucidating further the biosynthetic pathway for sulphoquinovose.

Solubilization and studies of cereal lipases

Abstract: accumulation. The induction of b-ketoacyl-ACP synthase 1 and 11 is similar to that of ACP (Slabas et ul., 1987) and enoyl-ACP reductase (Slabas et ul., 1986) from oilseed rape. In contrast, oilseed rape acetyl-CoA carboxylase shows a rapid loss of activity after fatty acid accumulation is complete (Turnham & Northcote, 1984). Details of a typical purification of P-ketoacyl-ACP synthase 1 from oilseed rape are given in Table 1. The key to the successful purification of P-ketoacyl-ACP synthase I was the use of f.p.1.c. Mono Q ion-exchange chromatography at pH 7.5 and pH 9.0. From 8500 mg of starting protein we were able to obtain 8 pg of homogeneous P-ketoacyl-ACP synthase 1. The low recovery of enzyme activity is an indication of the instability of P-ketoacyl-ACP synthase 1 during purification. The purified enzyme was also very unstable. The native M, values of P-ketoacyl-ACP synthase I and 11 were determined by gel filtration chromatography. The average hi', value of P-ketoacyl-ACP synthase I was 86 700 and for P-ketoacyl-ACP synthase 11 the average M , value was determined to be 87800. The subunit M, value of P-ketoacyl-ACP synthase 1 determined by SDS/polyacrylamide-gel electrophoresis was 43 000, suggesting that the enzyme is a homodimer. The native values reported here are very similar to those obtained for the P-ketoacyl-ACP synthase I and 11 from E. coli, i t . 80 000 and 85 000, respectively (Garwin et ul., 1980). The E. coli enzymes are also homodimeric. P-Ketoacyl-ACP synthase I co-purified with synthase I1 through ammonium sulphate fractionation and hydroxyapatite chromatography and the enzymes were only partially separated by gel filtration chromatography. However, the enzymes could be separated almost completely by f.p.1.c. Mono Q ion-exchange chromatography. Thus, we have been able to show that the two activities reside in different proteins.

Interferon-γ: an overview of signals, mechanisms and functions

Abstract: Interferon-gamma (IFN-gamma) coordinates a diverse array of cellular programs through transcriptional regulation of immunologically relevant genes. This article reviews the current understanding of IFN-gamma ligand, receptor, signal transduction, and cellular effects with a focus on macrophage responses and to a lesser extent, responses from other cell types that influence macrophage function during infection. The current model for IFN-gamma signal transduction is discussed, as well as signal regulation and factors conferring signal specificity. Cellular effects of IFN-gamma are described, including up-regulation of pathogen recognition, antigen processing and presentation, the antiviral state, inhibition of cellular proliferation and effects on apoptosis, activation of microbicidal effector functions, immunomodulation, and leukocyte trafficking. In addition, integration of signaling and response with other cytokines and pathogen-associated molecular patterns, such as tumor necrosis factor-alpha, interleukin-4, type I IFNs, and lipopolysaccharide are discussed.

Microalgal triacylglycerols as feedstocks for biofuel production: perspectives and advances

Abstract: Microalgae represent an exceptionally diverse but highly specialized group of micro-organisms adapted to various ecological habitats. Many microalgae have the ability to produce substantial amounts (e.g. 20-50% dry cell weight) of triacylglycerols (TAG) as a storage lipid under photo-oxidative stress or other adverse environmental conditions. Fatty acids, the building blocks for TAGs and all other cellular lipids, are synthesized in the chloroplast using a single set of enzymes, of which acetyl CoA carboxylase (ACCase) is key in regulating fatty acid synthesis rates. However, the expression of genes involved in fatty acid synthesis is poorly understood in microalgae. Synthesis and sequestration of TAG into cytosolic lipid bodies appear to be a protective mechanism by which algal cells cope with stress conditions, but little is known about regulation of TAG formation at the molecular and cellular level. While the concept of using microalgae as an alternative and renewable source of lipid-rich biomass feedstock for biofuels has been explored over the past few decades, a scalable, commercially viable system has yet to emerge. Today, the production of algal oil is primarily confined to high-value specialty oils with nutritional value, rather than commodity oils for biofuel. This review provides a brief summary of the current knowledge on oleaginous algae and their fatty acid and TAG biosynthesis, algal model systems and genomic approaches to a better understanding of TAG production, and a historical perspective and path forward for microalgae-based biofuel research and commercialization.