scholarly journals Distribution of the mevalonate and glyceraldehyde phosphate/pyruvate pathways for isoprenoid biosynthesis in unicellular algae and the cyanobacterium Synechocystis PCC 6714

1998 ◽  
Vol 333 (2) ◽  
pp. 381-388 ◽  
Author(s):  
Andrea DISCH ◽  
Jörg SCHWENDER ◽  
Christian MÜLLER ◽  
Hartmut K. LICHTENTHALER ◽  
Michel ROHMER
Keyword(s):  
Higher Plants ◽  
Scenedesmus Obliquus ◽  
Growth Conditions ◽  
Isoprenoid Biosynthesis ◽  
Mva Pathway ◽  
Feeding Experiments ◽  
Ochromonas Danica ◽  
Unicellular Algae ◽  
Β Carotene ◽  
Synechocystis Pcc 6714

Isopentenyl diphosphate, the universal isoprenoid precursor, can be produced by two different biosynthetic routes: either via the acetate/mevalonate (MVA) pathway, or via the more recently identified MVA-independent glyceraldehyde phosphate/pyruvate pathway. These two pathways are easily differentiated by incorporation of [1-13C]glucose and analysis of the resulting labelling patterns found in the isoprenoids. This method was successfully applied to several unicellular algae raised under heterotrophic growth conditions and allowed for the identification of the pathways that were utilized for isoprenoid biosynthesis. All isoprenoids examined (sterols, phytol, carotenoids) of the green algae Chlorella fusca and Chlamydomonas reinhardtii were synthesized via the GAP/pyruvate pathway, as in another previously investigated green alga, Scenedesmus obliquus, which was also shown in this study to synthesize ubiquinone by the same MVA-independent route. In the red alga Cyanidium caldarium and in the Chrysophyte Ochromonas danica a clear dichotomy was observed: as in higher plants, sterols were formed via the MVA route, whereas chloroplast isoprenoids (phytol in Cy. caldariumand O. danica and β-carotene in O. danica) were synthesized via the GAP/pyruvate route. In contrast, the Euglenophyte Euglena gracilis synthesized ergosterol, as well as phytol, via the acetate/MVA route. Similar feeding experiments were performed with the cyanobacterium SynechocystisPCC 6714 using [1-13C]- and [6-13C]-glucose. The two isoprenoids examined, phytol and β-carotene, were shown to have the typical labelling pattern derived from the GAP/pyruvate route.

scholarly journals Growth-uncoupled isoprenoid synthesis in Rhodobacter sphaeroides

2020 ◽  
Author(s):  
Enrico Orsi ◽  
Ioannis Mougiakos ◽  
Wilbert Post ◽  
Jules Beekwilder ◽  
Marco Dompè ◽  
...  
Keyword(s):  
Rhodobacter Sphaeroides ◽  
Metabolic Flux ◽  
Flux Ratio ◽  
Growth Conditions ◽  
Isoprenoid Biosynthesis ◽  
Microbial Cell ◽  
Cell Factory ◽  
Specific Product ◽  
Mva Pathway ◽  
Cell Factories

Abstract Background Microbial cell factories are usually engineered and employed for cultivations that combine product synthesis with growth. Such a strategy inevitably invests part of the substrate pool towards the generation of biomass and cellular maintenance. Hence, engineering strains for the formation of a specific product under non-growth conditions would allow to reach higher product yields. In this respect, isoprenoid biosynthesis represents an extensively studied example of growth-coupled synthesis with rather unexplored potential for growth-independent production. Rhodobacter sphaeroides is a model bacterium for isoprenoid biosynthesis, either via the native 2-methyl-D-erythritol 4-phosphate (MEP) pathway or the heterologous mevalonate (MVA) pathway, and for poly-β-hydroxybutyrate (PHB) biosynthesis. Results This study investigates the use of this bacterium for growth-independent production of isoprenoids, with amorpha-4,11-diene as reporter molecule. For this purpose, we employed the recently developed Cas9-based genome editing tool for R. sphaeroides to rapidly construct single and double deletion mutant strains of the MEP and PHB pathways, and we subsequently transformed the strains with the amorphadiene producing plasmid. Furthermore, we employed 13 C-metabolic flux ratio analysis to monitor the changes in the isoprenoid metabolic fluxes under different cultivation conditions. We demonstrated that active flux via both isoprenoid pathways while inactivating PHB synthesis maximizes growth-coupled isoprenoid synthesis. On the other hand, the strain that showed the highest growth-independent isoprenoid yield and productivity, combined the plasmid-based heterologous expression of the orthogonal MVA pathway with the inactivation of the native MEP and PHB production pathways. Conclusions Apart from proposing a microbial cell factory for growth-independent isoprenoid synthesis, this work provides novel insights about the interaction of MEP and MVA pathways under different growth-conditions.

scholarly journals Growth-uncoupled isoprenoid synthesis in Rhodobacter sphaeroides

2020 ◽  
Author(s):  
Enrico Orsi ◽  
Ioannis Mougiakos ◽  
Wilbert Post ◽  
Jules Beekwilder ◽  
Marco Dompè ◽  
...  
Keyword(s):  
Rhodobacter Sphaeroides ◽  
Metabolic Flux ◽  
Flux Ratio ◽  
Growth Conditions ◽  
Isoprenoid Biosynthesis ◽  
Microbial Cell ◽  
Cell Factory ◽  
Specific Product ◽  
Mva Pathway ◽  
Cell Factories

Abstract Background: Microbial cell factories are usually engineered and employed for cultivations that combine product synthesis with growth. Such a strategy inevitably invests part of the substrate pool towards the generation of biomass and cellular maintenance. Hence, engineering strains for the formation of a specific product under non-growth conditions would allow to reach higher product yields. In this respect, isoprenoid biosynthesis represents an extensively studied example of growth-coupled synthesis with rather unexplored potential for growth-independent production. Rhodobacter sphaeroides is a model bacterium for isoprenoid biosynthesis, either via the native 2-methyl-D-erythritol 4-phosphate (MEP) pathway or the heterologous mevalonate (MVA) pathway, and for poly-β-hydroxybutyrate (PHB) biosynthesis.Results: This study investigates the use of this bacterium for growth-independent production of isoprenoids, with amorpha-4,11-diene as reporter molecule. For this purpose, we employed the recently developed Cas9-based genome editing tool for R. sphaeroides to rapidly construct single and double deletion mutant strains of the MEP and PHB pathways, and we subsequently transformed the strains with the amorphadiene producing plasmid. Furthermore, we employed 13C-metabolic flux ratio analysis to monitor the changes in the isoprenoid metabolic fluxes under different cultivation conditions. We demonstrated that active flux via both isoprenoid pathways while inactivating PHB synthesis maximizes growth-coupled isoprenoid synthesis. On the other hand, the strain that showed the highest growth-independent isoprenoid yield and productivity, combined the plasmid-based heterologous expression of the orthogonal MVA pathway with the inactivation of the native MEP and PHB production pathways.Conclusions: Apart from proposing a microbial cell factory for growth-independent isoprenoid synthesis, this work provides novel insights about the interaction of MEP and MVA pathways under different growth-conditions.

scholarly journals The Coordinated Upregulated Expression of Genes Involved in MEP, Chlorophyll, Carotenoid and Tocopherol Pathways, Mirrored the Corresponding Metabolite Contents in Rice Leaves during De-Etiolation

Plants ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1456
Author(s):  
Xin Jin ◽  
Can Baysal ◽  
Margit Drapal ◽  
Yanmin Sheng ◽  
Xin Huang ◽  
...  
Keyword(s):  
Higher Plants ◽  
Expression Patterns ◽  
Regulatory Elements ◽  
Isoprenoid Biosynthesis ◽  
Promoter Regions ◽  
Expression Of Genes ◽  
Coordinated Expression ◽  
Rice Leaves ◽  
Mechanistic Basis ◽  
Phosphate Synthase

Light is an essential regulator of many developmental processes in higher plants. We investigated the effect of 4-hydroxy-3-methylbut-2-enyl diphosphate reductase 1/2 genes (OsHDR1/2) and isopentenyl diphosphate isomerase 1/2 genes (OsIPPI1/2) on the biosynthesis of chlorophylls, carotenoids, and phytosterols in 14-day-old etiolated rice (Oyza sativa L.) leaves during de-etiolation. However, little is known about the effect of isoprenoid biosynthesis genes on the corresponding metabolites during the de-etiolation of etiolated rice leaves. The results showed that the levels of α-tocopherol were significantly increased in de-etiolated rice leaves. Similar to 1-deoxy-D-xylulose-5-phosphate synthase 3 gene (OsDXS3), both OsDXS1 and OsDXS2 genes encode functional 1-deoxy-D-xylulose-5-phosphate synthase (DXS) activities. Their expression patterns and the synthesis of chlorophyll, carotenoid, and tocopherol metabolites suggested that OsDXS1 is responsible for the biosynthesis of plastidial isoprenoids in de-etiolated rice leaves. The expression analysis of isoprenoid biosynthesis genes revealed that the coordinated expression of the MEP (2-C-methyl-D-erythritol 4-phosphate) pathway, chlorophyll, carotenoid, and tocopherol pathway genes mirrored the changes in the levels of the corresponding metabolites during de-etiolation. The underpinning mechanistic basis of coordinated light-upregulated gene expression was elucidated during the de-etiolation process, specifically the role of light-responsive cis-regulatory motifs in the promoter region of these genes. In silico promoter analysis showed that the light-responsive cis-regulatory elements presented in all the promoter regions of each light-upregulated gene, providing an important link between observed phenotype during de-etiolation and the molecular machinery controlling expression of these genes.

scholarly journals Intermediates of tocopherol biosynthesis in the unicellular alga Scenedesmus obliquus. The presence of three isomeric methylphytylbenzoquinones

1987 ◽  
Vol 242 (2) ◽  
pp. 367-373 ◽  
Author(s):  
A Henry ◽  
R Powls ◽  
J F Pennock
Keyword(s):  
Carbon Atom ◽  
Hydroxy Group ◽  
Scenedesmus Obliquus ◽  
Alpha Tocopherol ◽  
Unicellular Alga ◽  
Unicellular Algae

Three isomers of methylphytylbenzoquinone have been isolated from lipids of the unicellular alga Scenedesmus obliquus, the most abundant being 2-methyl-6-phytylbenzoquinone (65% of the total). The 2-methyl-3-phytyl and 2-methyl-5-phytyl isomers amounted to 8 and 27% respectively. Previously problems have been encountered in the separation of the 3-phytyl and the 6-phytyl isomers, but in the present study it was found that they separated readily as quinols. Phytyl plastoquinone was also found and the relevance of these compounds to the biosynthesis of alpha-tocopherol is discussed. As well as phylloquinone, a hydroxyphylloquinone was detected, and studies indicated that it is the 5′ carbon atom to which the hydroxy group is attached. Such a compound has been found by workers using other unicellular algae.

The Non-Mevalonate Isoprenoid Biosynthesis of Plants as a Test System for New Herbicides and Drugs against Pathogenic Bacteria and the Malaria Parasite

2000 ◽  
Vol 55 (5-6) ◽  
pp. 305-313 ◽  
Author(s):  
Hartmut K. Lichtenthaler ◽  
Johannes Zeidler ◽  
Jörg Schwender ◽  
Christian Müller
Keyword(s):  
Malaria Parasite ◽  
Pathogenic Bacteria ◽  
Higher Plants ◽  
Test System ◽  
Isoprenoid Biosynthesis ◽  
Mep Pathway ◽  
New Drugs ◽  
Security Measures ◽  
Isoprenoid Pathway ◽  
Parasite Plasmodium Falciparum

Higher plants and several photosynthetic algae contain the plastidic 1-deoxy-ᴅ-xylulose 5- phosphate / 2-C-methyl-ᴅ-erythritol 4-phosphate pathway (DOXP/MEP pathway) for isoprenoid biosynthesis. The first four enzymes and their genes are known of this novel pathway. All of the ca. 10 enzymes of this isoprenoid pathway are potential targets for new classes of herbicides. Since the DOXP/MEP pathway also occurs in several pathogenic bacteria, such as Mycobacterium tuberculosis, and in the malaria parasite Plasmodium falciparum, all inhibitors and potential herbicides of the DOXP/MEP pathway in plants are also potential drugs against pathogenic bacteria and the malaria parasite. Plants with their easily to handle DOXP/MEP-pathway are thus very suitable test-systems also for new drugs against pathogenic bacteria and the malaria parasite as no particular security measures are required. In fact, the antibiotic herbicide fosmidomycin specifically inhibited not only the DOXP reductoisomerase in plants, but also that in bacteria and in the parasite P. falciparum, and cures malaria-infected mice. This is the first successful application of a herbicide of the novel isoprenoid pathway as a possible drug against malaria.

Biosynthese von p-Hydroxybenzoesäure und anderer Benzoesäuren in höheren Pflanzen

1964 ◽  
Vol 19 (5) ◽  
pp. 398-405 ◽  
Author(s):  
M. H. Zenk ◽  
G. Müller
Keyword(s):  
Benzoic Acid ◽  
Protocatechuic Acid ◽  
Higher Plants ◽  
Side Chain ◽  
Hydroxybenzoic Acid ◽  
Benzoic Acids ◽  
Coumaric Acid ◽  
Cinnamic Acids ◽  
Feeding Experiments ◽  
Catalpa Ovata

Feeding experiments with glucose- (2-14C), phenylalanine- (3-14C), tyrosine- (3-14C) and p-coumaric acid- (3-14C) showed that the latter three substances are incorporated in good yields into p-hydroxybenzoic acid in leaves of Catalpa ovata. Kinetic experiments showed that p-hydroxybenzoic acid is formed from phenylalanine via p-coumaric acid and the subsequent β-oxidation of the side chain. p-Hydroxybenzoic acid can also be synthetised by hydroxylation of benzoic acid, but this does not seem to be the biosynthetic route in Catalpa.Phenylalanine- (3-14C) is also incorporated into benzoic acid, protocatechuic acid, and vanillic acid by different plants; the radioactivity of the β-C atom of the amino acid was found in each case to be located in the carboxyl group of the C6 — C1 acid. This suggests that in higher plants the benzoic acids are formed from the corresponding cinnamic acids via β-oxidation.

The pigments of Artemia

1960 ◽  
Vol 152 (946) ◽  
pp. 118-136 ◽  
Keyword(s):  
Brine Shrimp ◽  
Phaeodactylum Tricornutum ◽  
Artemia Salina ◽  
Dunaliella Tertiolecta ◽  
Carotenoid Pigment ◽  
Algal Food ◽  
Feeding Experiments ◽  
Unicellular Algae ◽  
Bright Orange ◽  
Egg Shells

The eggs of the brine shrimp, Artemia salina , vary in colour from pale cream to dark brown. This variation is due to different amounts of haematin in the egg shells. Nauplii of Artemia are bright orange in colour owing to a carotenoid pigment, esterified astaxanthin. The same carotenoid is present in the eggs. Adult Artemia which has been reared on bakers’ yeast, in which we found no carotenoids, contains only a small amount of astaxanthin ester, presumably derived from that present in the egg. The carotenoids of the unicellular algae Dunaliella tertiolecta and Phaeodactylum tricornutum have been examined as a preliminary to feeding experiments with Artemia . The carotenoids identified from Dunaliella were β -carotene, γ -carotene, a carotene oxide, lutein, violaxanthin, and neoxanthin; those from Phaeodactylum were β -carotene, diadinoxanthin, fucoxanthin and neofucoxanthin. Adult Artemia reared on Dunaliella retains varying amounts of all the algal carotenoids, and in addition changes some of them to astaxanthin which becomes esterified and is quantitatively the most abundant carotenoid in the animal. A keto-carotenoid has been found in Artemia examined soon after being fed on Dunaliella . Artemia fed on Phaeodactylum retains all the algal xanthophylls to some extent. No β -carotene was found in the animals; a large amount of a keto-carotenoid was found, as well as astaxanthin. There is evidence that β -carotene in the algal food is the precursor of astaxanthin found in the adult Artemia and that the transformation proceeds through the keto-carotenoids.

scholarly journals Isoprenoid biosynthesis via 1-deoxy-D-xylulose 5-phosphate/2-C-methyl-D-erythritol 4-phosphate (DOXP/MEP) pathway.

2001 ◽  
Vol 48 (3) ◽  
pp. 663-672 ◽  
Author(s):  
M Wanke ◽  
K Skorupinska-Tudek ◽  
E Swiezewska
Keyword(s):  
Mevalonate Pathway ◽  
Higher Plants ◽  
Developmental State ◽  
Isoprenoid Biosynthesis ◽  
Mep Pathway ◽  
The Novel ◽  
Dimethylallyl Diphosphate ◽  
Parasite Plasmodium ◽  
Parasite Plasmodium Falciparum ◽  
Mixed Origin

Higher plants, several algae, bacteria, some strains of Streptomyces and possibly malaria parasite Plasmodium falciparum contain the novel, plastidic DOXP/MEP pathway for isoprenoid biosynthesis. This pathway, alternative with respect to the classical mevalonate pathway, starts with condensation of pyruvate and glyceraldehyde-3-phosphate which yields 1-deoxy-D-xylulose 5-phosphate (DOXP); the latter product can be converted to isopentenyl diphosphate (IPP) and eventually to isoprenoids or thiamine and pyridoxal. Subsequent reactions of this pathway involve transformation of DOXP to 2-C-methyl-D-erythritol 4-phosphate (MEP) which after condensation with CTP forms 4-diphosphocytidyl-2-amethyl-D-erythritol (CDP-ME). Then CDP-ME is phosphorylated to 4-diphosphocytidyl-2-amethyl-D-erythritol 2-phosphate (CDP-ME2P) and to 2-C-methyl-D-erythritol-2,4-cyclodiphosphate (ME-2,4cPP) which is the last known intermediate of the DOXP/MEP pathway. For- mation of IPP and dimethylallyl diphosphate (DMAPP) from ME-2,4cPP still requires clarification. This novel pathway appears to be involved in biosynthesis of carotenoids, phytol (side chain of chlorophylls), isoprene, mono-, di-, tetraterpenes and plastoquinone whereas the mevalonate pathway is responsible for formation of sterols, sesquiterpenes and triterpenes. Several isoprenoids were found to be of mixed origin suggesting that some exchange and/or cooperation exists between these two pathways of different biosynthetic origin. Contradictory results described below could indicate that these two pathways are operating under different physiological conditions of the cell and are dependent on the developmental state of plastids.

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