Engineering a platform for photosynthetic isoprene production in cyanobacteria, using Synechocystis as the model organism

Lindberg, Pia; Park, Sungsoon; Melis, Anastasios

doi:10.1016/j.ymben.2009.10.001

Cited by 547 publications

(406 citation statements)

References 42 publications

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“…MED134 (6). The drop in PR content during the stationary phase may be in part due to the promoter we selected, because it is known that psbA2 expression and transcript stability are light-intensity dependent (68)(69)(70)(71)(72)(73), and so is heterologous expression driven by this promoter (74,75). It is worth noting that several of the recently discovered proteorhodopsins in 'chemotrophic', often psychrophilic, bacteria only contribute detectably to growth rate or cell yield when the cells are stressed and/or in stationary phase (6,(14)(15)(16)(18)(19)(20).…”

Section: Discussionmentioning

confidence: 99%

Expression of holo-proteorhodopsin in Synechocystis sp. PCC 6803

Chen

Steen

Dekker

et al. 2016

Metabolic Engineering

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Retinal-based photosynthesis may contribute to the free energy conversion needed for growth of an organism carrying out oxygenic photosynthesis, like a cyanobacterium. After optimization, this may even enhance the overall efficiency of phototrophic growth of such organisms in sustainability applications. As a first step towards this, we here report on functional expression of the archetype proteorhodopsin in Synechocystis sp. PCC 6803. Upon use of the moderate-strength psbA2 promoter, holo-proteorhodopsin is expressed in this cyanobacterium, at a level of up to 10 5 molecules per cell, presumably in a hexameric quaternary structure, and with approximately equal distribution (on a protein-content basis) over the thylakoid and the cytoplasmic membrane fraction. These results also demonstrate that Synechocystis sp. PCC 6803 has the capacity to synthesize alltrans-retinal. Expressing a substantial amount of a heterologous opsin membrane protein causes a substantial growth retardation Synechocystis, as is clear from a strain expressing PROPS, a nonpumping mutant derivative of proteorhodopsin. Relative to this latter strain, proteorhodopsin expression, however, measurably stimulates its growth.

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Section: Discussionmentioning

confidence: 99%

Expression of holo-proteorhodopsin in Synechocystis sp. PCC 6803

Chen

Steen

Dekker

et al. 2016

Metabolic Engineering

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“…The main energy storage molecule in cyanobacteria is glycogen (Lindberg et al, 2010;Suzuki et al, 2010), which can be determined by assaying the amount of Glc following acid hydrolysis. Glycogen levels in the wild type and the COX/Cyd mutant cultured under 12-h high-light/dark square-wave cycles (Fig.…”

Section: Glycogen Measurements In the Wild Type And The Thylakoid Termentioning

confidence: 99%

Thylakoid Terminal Oxidases Are Essential for the CyanobacteriumSynechocystissp. PCC 6803 to Survive Rapidly Changing Light Intensities

et al. 2013

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Cyanobacteria perform photosynthesis and respiration in the thylakoid membrane, suggesting that the two processes are interlinked. However, the role of the respiratory electron transfer chain under natural environmental conditions has not been established. Through targeted gene disruption, mutants of Synechocystis sp. PCC 6803 were generated that lacked combinations of the three terminal oxidases: the thylakoid membrane-localized cytochrome c oxidase (COX) and quinol oxidase (Cyd) and the cytoplasmic membrane-localized alternative respiratory terminal oxidase. All strains demonstrated similar growth under continuous moderate or high light or 12-h moderate-light/dark square-wave cycles. However, under 12-h high-light/dark square-wave cycles, the COX/Cyd mutant displayed impaired growth and was completely photobleached after approximately 2 d. In contrast, use of sinusoidal light/dark cycles to simulate natural diurnal conditions resulted in little photobleaching, although growth was slower. Under high-light/dark square-wave cycles, the COX/Cyd mutant suffered a significant loss of photosynthetic efficiency during dark periods, a greater level of oxidative stress, and reduced glycogen degradation compared with the wild type. The mutant was susceptible to photoinhibition under pulsing but not constant light. These findings confirm a role for thylakoid-localized terminal oxidases in efficient dark respiration, reduction of oxidative stress, and accommodation of sudden light changes, demonstrating the strong selective pressure to maintain linked photosynthetic and respiratory electron chains within the thylakoid membrane. To our knowledge, this study is the first to report a phenotypic difference in growth between terminal oxidase mutants and wild-type cells and highlights the need to examine mutant phenotypes under a range of conditions.

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“…For several biofuel and chemical feedstock products, including ethanol, ethylene, isoprene, D-lactic acid, glucose, sucrose, isobutyraldehyde, and 1-butanol, the proof of principle of this approach has been provided (see references 3,8,10,24,25,28, and 39 and references therein). Ideally, this process would be carried out in such a way that the majority of the CO 2 that is fixed in the cyanobacterial CalvinBenson cycle is converted into the selected product.…”

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confidence: 99%

Engineering a Cyanobacterial Cell Factory for Production of Lactic Acid

Angermayr

Paszota

Hellingwerf

2012

Appl Environ Microbiol

165

144

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Metabolic engineering of microorganisms has become a versatile tool to facilitate production of bulk chemicals, fuels, etc. Accordingly, CO 2 has been exploited via cyanobacterial metabolism as a sustainable carbon source of biofuel and bioplastic precursors. Here we extended these observations by showing that integration of an ldh gene from Bacillus subtilis (encoding an L-lactate dehydrogenase) into the genome of Synechocystis sp. strain PCC6803 leads to L-lactic acid production, a phenotype which is shown to be stable for prolonged batch culturing. Coexpression of a heterologous soluble transhydrogenase leads to an even higher lactate production rate and yield (lactic acid accumulating up to a several-millimolar concentration in the extracellular medium) than those for the single ldh mutant. The expression of a transhydrogenase alone, however, appears to be harmful to the cells, and a mutant carrying such a gene is rapidly outcompeted by a revertant(s) with a wild-type growth phenotype. Furthermore, our results indicate that the introduction of a lactate dehydrogenase rescues this phenotype by preventing the reversion.C oncerns about shrinking supplies of fossil fuel and about climate change have been a strong incentive during the past decade for the development of sustainable alternatives to provide fuel and chemical feedstock. Because of these concerns, several different ways to produce solar biofuel have been proposed. Initially the proposed procedures were based on the conversion of agricultural crops into short-chain alcohols. This unfortunately results in a direct competition for resources between the energy sector and food supply. Therefore, second-generation types of processes were proposed, in which one aims at the conversion of the triglyceride fraction from, e.g., green algae into biodiesel, which relaxes the competition between energy and the food supply. Nevertheless, the conversion of CO 2 into biofuel, driven by solar energy, is still rather indirect even in this second-generation approach: CO 2 is first converted into the complex building blocks of biomass, from which subsequently the triglyceride fraction must be extracted and converted into biodiesel.For these reasons, a third-generation type of process has been proposed, in which CO 2 is converted into biofuel directly, accomplished by the introduction of genes encoding a metabolic-e.g., fermentative-pathway into a cyanobacterium, resulting in the emergence of a "photofermentative" chimera. For several biofuel and chemical feedstock products, including ethanol, ethylene, isoprene, D-lactic acid, glucose, sucrose, isobutyraldehyde, and 1-butanol, the proof of principle of this approach has been provided (see references 3,8,10,24,25,28, and 39 and references therein). Ideally, this process would be carried out in such a way that the majority of the CO 2 that is fixed in the cyanobacterial CalvinBenson cycle is converted into the selected product. This implies that the central metabolism in the selected chimera should be largely redirected tow...

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Engineering a platform for photosynthetic isoprene production in cyanobacteria, using Synechocystis as the model organism

Cited by 547 publications

References 42 publications

Expression of holo-proteorhodopsin in Synechocystis sp. PCC 6803

Expression of holo-proteorhodopsin in Synechocystis sp. PCC 6803

Thylakoid Terminal Oxidases Are Essential for the CyanobacteriumSynechocystissp. PCC 6803 to Survive Rapidly Changing Light Intensities

Engineering a Cyanobacterial Cell Factory for Production of Lactic Acid

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