C4 Intermediates Research Articles

100% selective cyclotrimerization of acetylene to benzene on Ag(111).

Benzene, a high-volume chemical, is produced from larger molecules by inefficient and environmentally harmful processes. Recent changes in hydrocarbon feedstocks from oil to gas motivate research into small molecule upgrading. For example, the cyclotrimerization of acetylene reaction has been demonstrated on Pd, Pd alloy, and Cu surfaces and catalysts, but they are not 100% selective to benzene. We discovered that acetylene can be converted to benzene with 100% selectivity on the Ag(111) surface. Our temperature programmed desorption experiments reveal a threshold acetylene surface coverage of ∼one monolayer, above which benzene is formed. Furthermore, additional layers of acetylene increase the amount of benzene produced while retaining 100% selectivity. Our scanning tunneling microscopy images show that acetylene prefers square packing on the Ag(111) surface at low coverages, which converts to hexagonal packing when acetylene multilayers are present. Within this denser layer, features consistent with the proposed C4 intermediates of the cyclotrimerization process are observed. Density functional theory calculations demonstrate that the barrier for forming the crucial C4 intermediate generally decreases as acetylene multilayers are formed because the multilayer interacts more strongly with the surface in the transition state than in the initial state. Given that acetylene desorbs from Ag(111) at ∼90 K, the C4 intermediate on the pathway to benzene must be formed below this temperature, implying that if Ag-based heterogeneous catalysts can be run at sufficiently high pressure and low enough temperature, efficient and selective trimerization of acetylene to benzene may be possible.

Breaking the C[sbnd]C bond of glucose on tungsten oxide-based catalysts in aqueous phase

In chemistry, selective activation of CC bonds enables the direct production of valuable chemicals from widely available and inexpensive natural materials but remains a fundamental challenge due to their kinetic inertness. The selective cleavage of CC bond in glucose by tungsten oxide-based catalysts in aqueous phase pioneers a path for converting cellulose biomass into valuable ethylene glycol. However, debates regarding the active phase and how it selectively breaks the C6 into C2 fragments have persisted for over a decade. In this study, we present a comprehensive mechanistic investigation by modeling three potential active phases, i.e. the reduced WO3-x surface, the dissolved tungstic acid, and tungsten bronze, in explicit solvent waters. By constrained molecular dynamics simulations, we have demonstrated that the low-coordinated W center can chelate with glucose, forming a metallacyclic complex with a 5-membered ring after the protonation of carbonyl group. The formation of 5-membered ring serves as the premise for the selectivity to C2 fragments via homolytic cleavage of CC bond. Furthermore, the reduced W5+ center is suggested to be crucial in facilitating the cleavage process by stabilizing the dissociated C4 intermediates via a redox process. In conclusion, we propose that the surface decoration of reduced and low-coordinated W sites can act as active heterogeneous catalysts for the selective conversion of cellulose in aqueous phase. These recent findings have the potential to provide valuable insights and strategies for CC bond activation in both biomass conversion and organic synthesis.

Soil organic carbon stock change following perennialization: a meta-analysis

Perennial crops replacing annual crops are drawing global attention because they harbor potential for sustainable biomass production and climate change mitigation through soil carbon sequestration. At present, it remains unclear how long perennial crops can sequester carbon in the soil and how soil carbon stock dynamics are influenced by climate, soil, and plant properties across the globe. This study presents a meta-analysis synthesizing 51 publications (351 observations at 77 sites) distributed over different pedo-climatic conditions to scrutinize the effect of perennialization on organic carbon accumulation in soil compared with two annual benchmark systems (i.e., monoculture and crop rotation). Results showed that perennial crops significantly increased soil organic carbon stock by 16.6% and 23.1% at 0–30 cm depth compared with monoculture and crop rotation, respectively. Shortly after establishment (< 5 years), perennial crops revealed a negative impact on soil organic carbon stock; however, long duration (> 10 years) of perennialization had a significant positive effect on soil organic carbon stock by 30% and 36.4% at 0–30 cm depth compared with monoculture and crop rotation, respectively. Compared with both annual systems, perennial crops significantly increased soil organic carbon stock regardless of their functional photosynthetic types (C3, C4, or C3-C4 intermediates) and vegetation type (woody or herbaceous). Among other factors, pH had a significant impact on soil organic carbon; however, the effect of soil textures showed no significant impact, possibly due to a lack of observations from each textural class and mixed pedoclimatic effects. Results also showed that time effect of perennialization revealed a sigmoidal increase of soil organic carbon stock until about 20 years; thereafter, the soil carbon stocks advanced towards a steady-state level. In conclusion, perennial crops increased soil organic carbon stock compared with annual systems; however, the time since conversion from annual to perennial system decisively impacted soil organic carbon stock changes.

Confinement-Enhanced Selective Oxidation of Lignin Derivatives to Formic Acid Over Fe-Cu/ZSM-5 Catalysts Under Mild Conditions.

Aqueous-phase oxidation by H2 O2 , known as the Fenton-type process, provides an attractive route to convert recalcitrant lignin derivatives to valuable chemicals under mild conditions. The development of this technology is, however, limited by the uncontrolled selectivity, resulting from the highly reactive nature of H2 O2 and the thermodynamically favored deep oxidation to form CO2 . This study demonstrated that formic acid could be produced with a high selectivity (up to 80.3 % at 313 K) from the Fenton-type oxidation of guaiacol and several other lignin derivatives over a bimetallic Fe-Cu catalyst supported on a ZSM-5 zeolite. Combined experimental and theoretical investigations unveiled that the micropores of the zeolite support, which contained active metal sites, preferred to adsorb C2 -C4 intermediates over formic acid because of its stronger dispersive interaction with the larger guest molecules. This confinement effect significantly suppressed the secondary oxidation of formic acid, accounting for the uniquely high formic acid selectivity over Fe-Cu/ZSM-5.

Biosynthesis of Poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) From Glucose by Escherichia coli Through Butyryl-CoA Formation Driven by Ccr-Emd Combination.

Poly[(R)-3-hydroxybutyrate-co-(R)-3-hydroxyhexanoate] [P(3HB-co-3HHx)] is a practical kind of bacterial polyhydroxyalkanoates (PHAs). A previous study has established an artificial pathway for the biosynthesis of P(3HB-co-3HHx) from structurally unrelated sugars in Ralstonia eutropha, in which crotonyl-CoA carboxylase/reductase (Ccr) and ethylmalonyl-CoA decarboxylase (Emd) are a key combination for generation of butyryl-CoA and the following chain elongation. This study focused on the installation of the artificial pathway into Escherichia coli. The recombinant strain of E. coli JM109 harboring 11 heterologous genes including Ccr and Emd produced P(3HB-co-3HHx) composed of 14 mol% 3HHx with 41 wt% of dry cellular weight from glucose. Further investigations revealed that the C6 monomer (R)-3HHx-CoA was not supplied by (R)-specific reduction of 3-oxohexanoyl-CoA but by (R)-specific hydration of 2-hexenoyl-CoA formed through reverse β-oxidation after the elongation from C4 to C6. While contribution of the reverse β-oxidation to the conversion of the C4 intermediates was very limited, crotonyl-CoA, a precursor of butyryl-CoA, was generated by dehydration of (R)-3HB-CoA. Several modifications previously reported for enhancement of bioproduction in E. coli were examined for the copolyester synthesis. Elimination of the global regulator Cra or PdhR as well as the block of acetate formation resulted in poor PHA synthesis. The strain lacking RNase G accumulated more PHA but with almost no 3HHx unit. Introduction of the phosphite oxidation system for regeneration of NADPH led to copolyester synthesis with the higher cellular content and higher 3HHx composition by two-stage cultivation with phosphite than those in the absence of phosphite.

Metabolic profiles in C3, C3-C4 intermediate, C4-like, and C4 species in the genus Flaveria.

C4 photosynthesis concentrates CO2 around Rubisco in the bundle sheath, favouring carboxylation over oxygenation and decreasing photorespiration. This complex trait evolved independently in >60 angiosperm lineages. Its evolution can be investigated in genera such as Flaveria (Asteraceae) that contain species representing intermediate stages between C3 and C4 photosynthesis. Previous studies have indicated that the first major change in metabolism probably involved relocation of glycine decarboxylase and photorespiratory CO2 release to the bundle sheath and establishment of intercellular shuttles to maintain nitrogen stoichiometry. This was followed by selection for a CO2-concentrating cycle between phosphoenolpyruvate carboxylase in the mesophyll and decarboxylases in the bundle sheath, and relocation of Rubisco to the latter. We have profiled 52 metabolites in nine Flaveria species and analysed 13CO2 labelling patterns for four species. Our results point to operation of multiple shuttles, including movement of aspartate in C3–C4 intermediates and a switch towards a malate/pyruvate shuttle in C4-like species. The malate/pyruvate shuttle increases from C4-like to complete C4 species, accompanied by a rise in ancillary organic acid pools. Our findings support current models and uncover further modifications of metabolism along the evolutionary path to C4 photosynthesis in the genus Flaveria.

Exploring combustion chemistry of 1‐pentene: Flow reactor pyrolysis at various pressures and development of a detailed combustion model

AbstractThe pyrolysis of 1‐pentene was investigated in a flow reactor over temperatures of 900–1300 K and pressures of 0.04 and 1 atm using synchrotron vacuum ultraviolet photoionization mass spectrometry (SVUV‐PIMS). Pyrolysis products, especially radicals and cyclic compounds, were identified, and their mole fraction profiles were quantified. A detailed kinetic model of 1‐pentene combustion was developed and validated against the new data in this work. Modeling analysis including rate of production analysis and sensitivity analysis was performed to reveal the key pathways in the decomposition of 1‐pentene and the formation of pyrolysis products. Due to the presence of CC double bond in 1‐pentene molecule, the allylic CC bond dissociation reaction and the retro‐ene reaction play important roles in the formation of dominant pyrolysis products such as small alkenes. Besides, the formation of cyclic products including 1,3‐cyclopentadiene, fulvene, benzene, and toluene is dominated by the reactions of C3 intermediates with C2–C4 intermediates. The model was further validated against the previous data of 1‐pentene combustion, including the ignition delay times and laminar burning velocities over a wide range of pressures, temperatures, and equivalence ratios. Modeling analysis shows that C0–C3 reactions and fuel‐specific reactions are important for the ignition process of 1‐pentene, while C0–C2 reactions play significant roles in the laminar flame propagation of 1‐pentene.

Phenotypic plasticity of leaf anatomical traits helps to explain gas-exchange response to water shortage in grasses of different photosynthetic types

C3 and C4 plants, as their intermediates, respond differently to short-term changes in environmental conditions. This difference is linked to contrasting levels of phenotypic plasticity and photosynthetic apparatus specialization. Phenotypic plasticity is an underexplored topic although its understanding is crucial to predict plant behaviour in future climatic scenarios. In this research, the phenotypic plasticity of anatomical traits and its influence to carbon uptake efficiency was studied in plants with different photosynthetic types, under contrasting water regimes. Oryza sativa cvs. Soberana (drought-sensitive) and Douradao (drought-tolerant) (C3), Homolepis isocalycia (C3 proto-Kranz) and Andropogon gayanus (C4), grown at three water treatments (100, 75 and 50% of substrate water holding capacity), were phenotyped for leaf anatomy and gas-exchange parameters. The results showed that plasticity trends indicated different strategies between O. sativa cultivars to deal with water shortage, explaining their classification as drought-sensitive or tolerant. We also mapped typical characteristics of C3–C4 intermediate plant, H. isocalycia, mainly in the ratio mesophyll:bundle sheath cells and hypothesize how it may influence photosynthesis. Finally, we have confirmed previous claims that C4 carbon uptake advantages may be limited under severe drought conditions, as A. gayanus have drastically reduced its photosynthetic rates at lower water levels. By studying C3–C4 intermediates, this study may also be a starting point to unravel the trade-offs of anatomical changes during the evolutionary process from C3 to C4 photosynthesis, and also improve the understanding of their impact in carbon uptake in different water conditions.

Synthesis of C4 Olefins from Acetylene over Supported Copper Catalysts

AbstractConversion of acetylene to C4 olefins enables on‐purpose synthesis of C4 olefins using abundant natural gas resource due to acetylene can be synthesized from methane or ethane. In this study, Cu‐based catalysts supported on SiO2, ZSM‐5 and mordenite (MOR) were tested for acetylene hydrodimerization. Effect of promoters including Ru, Ga, Ag and Pd on the acetylene conversion and C4 olefins selectivity are investigated. The structures of these catalysts were characterized by TEM, XRD, BET, H2‐TPR, NH3‐TPD and XPS measurements. Results showed that temperature, pressure and hydrogen partial pressure had positive effect on acetylene conversion. In the presence of Ru promotor, 100 % acetylene conversion and 87 % selectivity to C4 olefins were achieved over Cu/MOR catalyst under conditions of 0.1 MPa and 200 °C. The mechanistic aspect of acetylene hydrodimerization was elucidated where the C4 intermediate was formed by the reaction between [Cu]−C2H2 π‐complex and [Cu]−C2H2 σ‐complex which were transformed to 1,3‐butadiene and butene.

Identifying the roles of acid–base sites in formation pathways of tolualdehydes from acetaldehyde over MgO-based catalysts

Basic (M–O)-type centers convert C4 intermediates to renewable xylene analogs and proximal acid sites tune isomeric selectivity.

Selective Oxidation of n-Butane over Vanadium Phosphate Based Catalysts: Reaction Network and Kinetic Analysis

The reaction network of n-butane selective oxidation was comparatively analyzed over a novel αII-V0.8W0.2OPO4 orthophosphate and a reference vanadyl pyrophosphate (VPP) based catalyst via parameter field studies (T = 360–420 °C, x(n-butane) = 0.5–2.0%, x(O2) = 10–20%) as well as cofeed and pulse experiments with presumed reaction intermediates. For VPP the selectivity to the target product maleic anhydride (MAN) is particularly sensitive to the reaction temperature, the n-butane concentration, and the amount of cofed H2O, whereas the selectivity to MAN over the W-containing catalyst is almost independent of all reaction parameters. The roles of 1-butene, acetylene, furan, acetaldehyde, water, and 2,5-dihydrofuran are discussed. Pulsing of possible C4 intermediates indicates that their desorption from the catalyst surface is detrimental to MAN selectivity. Ethylene and acetylene may be formed from MAN. The consecutive reaction of acetylene with H2O can lead to acetic acid, whereas all other byproducts are ...

Tendencies towards a C4 Leaf: Quantitative Studies on Leaf Anatomy of Selected C3 and C4 Grasses

Grass family (Poaceae) is one of the largest families that represent monocots; containing species like Oryza sativa, Triticum aestivum, Zea mays, Eleusine coracana, Panicum miliaceum, Panicum sumatrense and Sorghum bicolor which are important to humans since they, directly or indirectly provide more than 3/4 of our food. C4 photosynthesis is a complex physiological adaptation, that is believed to have resulted from a series of anatomical and biochemical modifications to the ancestral C3 pathway. It is believed that C4 photosynthesis to have evolved in a stepwise manner nearly 22-24 times creating intermediates with different combinations of C4 -like components in grasses. C4 photosynthesis confers greater productivity than the C3 photosynthetic type in environments under high irradiance and high temperature. Accompanying higher photosynthetic rate C4 plants possess a vascular system in leaves with higher export rates which permit efficient translocation of photosynthates. In the current study, selected 11 grass species were subjected to analysis of leaf vascular system in relation to their C3 and C4 photosynthesis. The leaf vascular network in both C3 and C4 grasses consist of LLV, SLV and TV that are connected to longitudinal veins. Analysis carried out using the mature leaves of selected C3 and C4 grasses have shown that there are significant differences in anatomical features such as venation, stomata and bundle sheaths. The comparison of DLLV, DSLV and DTV of C3 and C4 grasses demonstrate that C4 grasses have a denser vascular pattern compared to C3 species (respective distances of C4 grasses were 71.74 μm, 11.85 μm and 81.4 μm). It has been found that these reduced distances between veins have led to significant low stomatal density among C4 grasses. Further, characteristic large bundle sheath structure of C4 species has been clearly seen from significantly high values obtained for BSD, ISD and OSD (respectively 10.4 μm, 4.32 μm and 7.15 μm). Furthermore, this screening has found Oryza nivara (a C3 species) having C4 like anatomical characters in leaf venation and bundle sheath structure. This species showed significantly low mean values for the distance between transverse veins (41.75 μm) resulting in significantly high total vein length per unit leaf area (81.77μm-1). The C4 tendency of its leaf anatomy was further supported by significantly large bundle sheaths, inner sheath and outer sheath distances (13.1 μm, 5.22 μm and 8.22 μm respectively). At the end of the analysis we were able to demonstrate significant differences of anatomical characters like venation, stomata and bundle sheaths in C3 and C4 plants. In addition, observations suggest characteristic leaf anatomy that is compatible with C4 anatomy may have evolved in Oryza nivara resulting it to be considered as C3- C4 intermediate.

C3 -C4 intermediates may be of hybrid origin - a reminder.

The currently favoured model of the evolution of C4 photosynthesis relies heavily on the interpretation of the broad phenotypic range of naturally growing C3 -C4 intermediates as proxies for evolutionary intermediate steps. On the other hand, C3 -C4 intermediates had earlier been interpreted as hybrids or hybrid derivates. By first comparing experimentally generated with naturally growing C3 -C4 intermediates, and second summarising either direct or circumstantial evidence for hybridisation in lineages comprising C3 , C4 and C3 -C4 intermediates, we conclude that a possible hybrid origin of C3 -C4 intermediates deserves careful examination. While we acknowledge that the current model of C4 photosynthesis evolution is clearly the best available, C3 -C4 intermediates of hybrid origin, if existing, should not be used for further analysis of this model. However, experimental C3 ×C4 hybrids potentially are excellent systems to analyse the genetic differences between C3 and C4 species and, also using segregating progeny, to study the relationship between individual photosynthetic traits and environmental factors.

Heterologous expression of the human Phosphoenol Pyruvate Carboxykinase (hPEPCK-M) improves hydrogen and ethanol synthesis in the Escherichia coli dcuD mutant when grown in a glycerol-based medium

The production of biodiesel has emerged as an alternative to fossil fuels. However, this industry generates glycerol as a by-product in such large quantities that it has become an environmental problem. The biotransformation of this excess glycerol into other renewable bio-energy sources, like H2 and ethanol, by microorganisms such as Escherichia coli is an interesting possibility that warrants investigation. In this work we hypothesized that the conversion of oxaloacetate (OAA) to phosphoenolpyruvate (PEP) could be improved by a controlled expression of the human mitochondrial GTP-dependent PEP carboxykinase. This heterologous expression was tested in several E. coli mutant backgrounds with increased availability of C4 intermediates. It was found that this metabolic rewiring improved the synthesis of the target products in several mutants, with the dcuD mutant being the most suitable background for hydrogen and ethanol specific productions and glycerol consumption. These factors increased by 2.46, 1.73 and 1.95 times, respectively, when compared to those obtained for the wild-type strain.

Broadening the spectrum of photosynthesis in the grass, Alloteropsis semialata.

This article comments on: Evolutionary implications of C3 -C4 intermediates in the grass Alloteropsis semialata.

A phylogenetic study of subtribe Otachyriinae (Poaceae, Panicoideae, Paspaleae)

The subtribe Otachyriinae was established, within tribe Paniceae, by Butzin in 1970, originally including 18 genera. Recent phylogenetic studies, based on molecular data, indicated that Otachyriinae belongs to tribe Paspaleae and is represented by the genera Anthaenantia, Hymenachne, Dallwatsonia, Otachyrium, species of Panicum sect. Laxa, Plagiantha, and Steinchisma. These taxa show a peculiar diversity in photosynthetic types, including C4 species, C3–C4 intermediates and C3 species. The internal evolutionary relationships among members of Otachyriinae have not yet been explored in detail. With the aim of providing new evidence to understand phylogenetic relationships of taxa included in Otachyriinae, 21 new sequences of the ndhF gene from 34 species were generated and analyzed using Bayesian inference and maximum parsimony (MP). The evolution of photosynthetic pathway was studied employing MP, Markov chain Monte Carlo, and maximum likelihood methods. Our results confirm that subtribe Otachyriinae is monophyletic. Within the subtribe, Anthaenanthia is a monophyletic C4 genus and is sister to the remaining species included in this treatment. Hymenachne and Panicum sect. Laxa, in its traditional concept, are polyphyletic; Dallwatsonia, P. auritum, and P. hemitomon are treated within Hymenachne, while three species of Panicum sect. Laxa are considered a new genus in the subtribe. The genus Otachyrium is paraphyletic, while all species of Steinchisma are recovered as a monophyletic group with high support. Finally, our data suggest that C4 and C3–C4 photosynthesis evolved just once each during the speciation of the subtribe from a C3 ancestor.

Triheptanoin reduces seizure susceptibility in a syndrome-specific mouse model of generalized epilepsy

Triheptanoin is a triglyceride containing heptanoate, an odd-chained medium fatty acid that is metabolized to produce propionyl-CoA and subsequently C4 intermediates of the citric acid cycle and therefore capable of anaplerosis. These metabolic products are believed to underlie triheptanoin's anticonvulsant effects in rodent seizure models. Here we investigate the anticonvulsive effects of oral triheptanoin in a syndrome-specific genetic mouse model of generalized epilepsy based on the GABA(A)γ2(R43Q) mutation. Mice were fed a diet supplemented with triheptanoin from weaning for three weeks prior to electrocortical recordings. Occurrence and durations of spike and wave discharges (SWDs) were measured. Triheptanoin did not alter body weight or basal blood glucose levels suggesting that it was well tolerated. Triheptanoin supplementation halved the time spent in seizures due to a reduction in both SWD occurrence and duration. An injection of insulin was used to reduce blood glucose, a metabolic stress known to precipitate seizures in the GABA(A)γ2(R43Q) mouse. The reduction in seizure count was also evident following insulin induced hypoglycemia with the triheptanoin treated group having significantly less SWDs than control animals under similar low blood glucose conditions. In summary, triheptanoin may be an effective and well tolerated dietary therapy for generalized epilepsy.

Molecular phylogenetics of Alternanthera (Gomphrenoideae, Amaranthaceae): resolving a complex taxonomic history caused by different interpretations of morphological characters in a lineage with C4 and C3-C4 intermediate species

Alternanthera (Amaranthaceae) is a diverse genus (80–200 species) largely restricted to the American Tropics. With Pedersenia and Tidestromia, it makes up the ‘Alternantheroid clade’ in Gomphrenoideae. Parsimony and Bayesian analyses of nucleotide sequences of nuclear (ITS) and plastid (rpl16, trnL-F) and morphological characters identify that the capitate stigma of Alternanthera is a synapomorpy within the Alternantheroids. Within Alternanthera, two major clades were resolved, both of which were marked by otherwise homoplasious characters of the gynoecium: Clade A [99% jackknife (JK); 1.0 posterior probability (PP)] with nine species and Clade B (60% JK; 0.98 PP) with 22 species. Four subclades (B1–B4), strongly supported statistically, were identified in Clade B. Previous subgeneric classifications of Alternanthera appear artificial in light of our new molecular phylogenetic analyses. Most major lineages are congruently resolved by nuclear and plastid data but some incongruence between the nrITS and plastid phylogenetic trees suggests hybridization may have played a role in the rampant speciation in Alternanthera. Whereas C4 photosynthesis appears to have evolved in a single clade, the position of A. littoralis var. maritima (C3) in this clade may be explained by hybrid speciation rather than a reversal from C4 to C3. All C3–C4 intermediates belong to a different clade that also contains C3 species, but species limits, including the widely studied A. tenella, are unclear. The clade including A. tenella and A. halimifolia contains most of the species endemic to the Galapagos whereas A. nesiotes, also endemic to the islands, is nested among widespread American taxa. This suggests that the Galapagos radiation of Alternanthera may have arisen from at least two independent colonization events followed by a subsequent radiation in the former lineage. © 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2012, 169, 493–517.

Impact of Malic Enzymes on Antibiotic and Triacylglycerol Production in Streptomyces coelicolor

In this paper, we have characterized two malic enzymes (ME), SCO2951 and SCO5261, from Streptomyces coelicolor and analyzed their role in antibiotic and triacylglycerol (TAG) production. Biochemical studies have demonstrated that Sco2951 and Sco5261 genes encode NAD(+)- and NADP(+)-dependent malic enzymes, respectively. Single or double mutants in the ME-encoding genes show no effect on growth rate compared to the parental M145 strain. However, the single Sco2951 and the double Sco2951 Sco5261 mutants display a strong reduction in the production of the polyketide antibiotic actinorhodin; additionally, the Sco2951 Sco5261 mutant shows a decrease in stored TAGs during exponential growth. The lower production of actinorhodin in the double mutant occurs as a consequence of a decrease in the expression of actII-ORF4, the transcriptional activator of the actinorhodin gene cluster. On the other hand, the reduced TAG accumulation is not due to reduced transcript levels of fatty acid biosynthetic genes nor to changes in the amount of the precursor acetyl coenzyme A (acetyl-CoA). This mutant accumulates intermediates of the tricarboxylic acid (TCA) cycle that could alter the regulation of the actinorhodin biosynthetic pathway, suggesting that MEs are important anaplerotic enzymes that redirect C4 intermediates from the TCA cycle to maintain secondary metabolism and TAG production in Streptomyces.

Structural and biochemical characterization of the C₃-C₄ intermediate Brassica gravinae and relatives, with particular reference to cellular distribution of Rubisco.

On the basis of its CO2 compensation concentration, Brassica gravinae Ten. has been reported to be a C3–C4 intermediate. This study investigated the structural and biochemical features of photosynthetic metabolism in B. gravinae. The cellular distribution of ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) was also examined in B. gravinae, B. napus L. (C3), Raphanus sativus L. (C3), and Diplotaxis tenuifolia (L.) DC. (C3–C4) by immunogold electron microscopy to elucidate Rubisco expression during the evolution from C3 to C3–C4 intermediate plants. The bundle sheath (BS) cells of B. gravinae contained centrifugally located chloroplasts as well as centripetally located chloroplasts and mitochondria. Glycine decarboxylase P-protein was localized in the BS mitochondria. Brassica gravinae had low C4 enzyme activities and high activities of Rubisco and photorespiratory enzymes, suggesting that it reduces photorespiratory CO2 loss by the glycine shuttle. In B. gravinae, the labelling density of Rubisco was higher in the mesophyll chloroplasts than in the BS chloroplasts. A similar cellular pattern was found in other Brassicaceae species. These data demonstrate that, during the evolution from C3 to C3–C4 intermediate plants, the intercellular pattern of Rubisco expression did not change greatly, although the amount of chloroplasts in the BS cells increased. It also appears that intracellular variation in Rubisco distribution may occur within the BS cells of B. gravinae.