Cyanobacteria are photosynthetic prokaryotes showing great promise as biocatalysts for the direct conversion of CO2 into fuels, chemicals, and other value-added products. Introduction of just a few ...heterologous genes can endow cyanobacteria with the ability to transform specific central metabolites into many end products. Recent engineering efforts have centered around harnessing the potential of these microbial biofactories for sustainable production of chemicals conventionally produced from fossil fuels. Here, we present an overview of the unique chemistry that cyanobacteria have been co-opted to perform. We highlight key lessons learned from these engineering efforts and discuss advantages and disadvantages of various approaches.
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•Host organisms need to tolerate outdoor conditions and abiotic stresses.•Robust, fast-growing cyanobacterial strains can be engineered as efficient hosts.•Engineered and ...well-characterized host strains should be made easily accessible.•Synthetic biology protocols and parts need to be standardized for non-model strains.•Metabolomics assisted and model-guided pathway engineering is an emerging field.
Cyanobacteria, a group of photoautotrophic prokaryotes, are attractive hosts for the sustainable production of chemicals from carbon dioxide and sunlight. However, the rates, yields, and titers have remained well below those needed for commercial deployment. We argue that the following areas will be central to the development of cyanobacterial cell factories: engineered and well-characterized host strains, model-guided pathway design, and advanced synthetic biology tools. Although several foundational studies report improved strain properties, translational research will be needed to develop engineered hosts and deploy them for metabolic engineering. Further, the recent developments in metabolic modeling and synthetic biology of cyanobacteria will enable nimble strategies for strain improvement with the complete cycle of design, build, test, and learn.
•Extremophilic micro-algae have a potential role in the biotechnology industry.•We consider extremes of temperature, light, CO2, pH, salt and metal in this review.•We present phylogenetic analysis of ...extremophilic microalgae.•We discuss organisms' adaptive mechanisms to tackle these stresses.•Physiology, metabolic engineering and molecular biology need further studies.
Micro-algae have potential as sustainable sources of energy and products and alternative mode of agriculture. However, their mass cultivation is challenging due to low survival under harsh outdoor conditions and competition from other, undesired, species. Extremophilic micro-algae have a role to play by virtue of their ability to grow under acidic or alkaline pH, high temperature, light, CO2 level and metal concentration. In this review, we provide several examples of potential biotechnological applications of extremophilic micro-algae and the ranges of tolerated extremes. We also discuss the adaptive mechanisms of tolerance to these extremes. Analysis of phylogenetic relationship of the reported extremophiles suggests certain groups of the Kingdom Protista to be more tolerant to extremophilic conditions than other taxa. While extremophilic microalgae are beginning to be explored, much needs to be done in terms of the physiology, molecular biology, metabolic engineering and outdoor cultivation trials before their true potential is realized.
SUMMARY Synechococcus elongatus PCC 11801 and 11802 are closely related cyanobacterial strains that are fast‐growing and tolerant to high light and temperature. These strains hold significant promise ...as chassis for photosynthetic production of chemicals from carbon dioxide. A detailed quantitative understanding of the central carbon pathways would be a reference for future metabolic engineering studies with these strains. We conducted isotopic non‐stationary 13C metabolic flux analysis to quantitively assess the metabolic potential of these two strains. This study highlights key similarities and differences in the central carbon flux distribution between these and other model/non‐model strains. The two strains demonstrated a higher Calvin–Benson–Bassham (CBB) cycle flux coupled with negligible flux through the oxidative pentose phosphate pathway and the photorespiratory pathway and lower anaplerosis fluxes under photoautotrophic conditions. Interestingly, PCC 11802 shows the highest CBB cycle and pyruvate kinase flux values among those reported in cyanobacteria. The unique tricarboxylic acid (TCA) cycle diversion in PCC 11801 makes it ideal for the large‐scale production of TCA cycle‐derived chemicals. Additionally, dynamic labeling transients were measured for intermediates of amino acid, nucleotide, and nucleotide sugar metabolism. Overall, this study provides the first detailed metabolic flux maps of S. elongatus PCC 11801 and 11802, which may aid metabolic engineering efforts in these strains.
Synechococcus elongatus PCC 11801 and 11802 are closely related cyanobacterial strains that are fast-growing and tolerant to high light and temperature. These strains hold significant promise as ...chassis for photosynthetic production of chemicals from carbon dioxide. A detailed quantitative understanding of the central carbon pathways would be a reference for future metabolic engineering studies with these strains. We conducted isotopic non-stationary
C metabolic flux analysis to quantitively assess the metabolic potential of these two strains. This study highlights key similarities and differences in the central carbon flux distribution between these and other model/non-model strains. The two strains demonstrated a higher Calvin-Benson-Bassham (CBB) cycle flux coupled with negligible flux through the oxidative pentose phosphate pathway and the photorespiratory pathway and lower anaplerosis fluxes under photoautotrophic conditions. Interestingly, PCC 11802 shows the highest CBB cycle and pyruvate kinase flux values among those reported in cyanobacteria. The unique tricarboxylic acid (TCA) cycle diversion in PCC 11801 makes it ideal for the large-scale production of TCA cycle-derived chemicals. Additionally, dynamic labeling transients were measured for intermediates of amino acid, nucleotide, and nucleotide sugar metabolism. Overall, this study provides the first detailed metabolic flux maps of S. elongatus PCC 11801 and 11802, which may aid metabolic engineering efforts in these strains.
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•Repertoire of industrially relevant cyanobacterial strains needs to be expanded.•Genome engineering tools and well-characterized synthetic biology parts are needed.•Systems biology ...knowledge bases will help in developing new strains.•Desirable industrial traits can be engineered in selected host strains.•Metabolic engineering strategies tailored for new strains will improve yields.
Cyanobacteria, a group of photosynthetic prokaryotes, can be engineered for direct conversion of carbon dioxide to value added products. Despite two decades of progress in the metabolic engineering of these photoautotrophs, the reported productivities are below those needed for commercialization. While much of this work has been performed with a handful of model cyanobacteria, new fast-growing and robust cyanobacterial strains may afford significant improvements in productivity. The focus now is on isolating and developing cyanobacteria that can be deployed as industrial strains for the production of a wide range of chemicals. Systems-level characterization, development of synthetic biology tools and improvement in photosynthetic efficiency would be an integral part of host engineering efforts. The knowledge base that exists for model cyanobacteria together with the systems biology data from newer strains will provide the foundation for the development of new hosts.
A key requirement for 13C Metabolic flux analysis (13C-MFA), a widely used technique to estimate intracellular metabolic fluxes, is an efficient method for the extraction of intermediate metabolites ...for analysis via liquid chromatography mass spectrometry (LC/MS). The 13C isotopic labeling results in further distribution of an already sparse pool of intermediate metabolites into isotopologues, each appearing as a separate chromatographic feature. We examined some of the reported solvent systems for the extraction of polar intracellular metabolites from three strains of cyanobacteria of the genus Synechococcus, viz., Synechococcus sp. PCC 7002, Synechococcus elongatus PCC 7942, and a newly isolated Synechococcus elongatus PCC 11801 (manuscript under review). High resolution-LC/MS was used to assess the relative abundance of the extracted metabolites. The different solvent systems used for extraction led to statistically significant changes in the extraction efficiency for a large number of metabolites. While a few hundred m/z features or potential metabolites were detected with different solvent systems, the abundance of over a quarter of all metabolites varied significantly from one solvent system to another. Further, the extraction methods were evaluated for a targeted set of metabolites that are important in 13C-MFA studies of photosynthetic organisms. While for the strain PCC 7002, the reported method using methanol-chloroform-water system gave satisfactory results, a mild base in the form of NH4OH had to be used in place of water to achieve adequate levels of extraction for PCC 7942 and PCC 11801. While minor changes in extraction solvent resulted in dramatic changes in the extraction efficiency of a number of compounds, certain metabolites such as amino acids and organic acids were adequately extracted in all the solvent systems tested. Overall, we present a new improved method for extraction using a methanol-chloroform-NH4OH system. Our method improves the extraction of polar compounds such as sugar phosphates, bisphosphates, that are central to 13C-MFA studies.
Cyanobacteria are attractive hosts that can be engineered for the photosynthetic production of fuels, fine chemicals, and proteins from CO
2
. Moreover, the responsiveness of these photoautotrophs ...towards different environmental signals, such as light, CO
2
, diurnal cycle, and metals make them potential hosts for the development of biosensors. However, engineering these hosts proves to be a challenging and lengthy process. Synthetic biology can make the process of biological engineering more predictable through the use of standardized biological parts that are well characterized and tools to assemble them. While significant progress has been made with model heterotrophic organisms, many of the parts and tools are not portable in cyanobacteria. Therefore, efforts are underway to develop and characterize parts derived from cyanobacteria. In this review, we discuss the reported parts and tools with the objective to develop cyanobacteria as cell factories or biosensors. We also discuss the issues related to characterization, tunability, portability, and the need to develop enabling technologies to engineer this “green” chassis.
Cyanobacteria are attractive candidates for photoautotrophic production of platform chemicals due to their inherent ability to utilize carbon dioxide as the sole carbon source. Metabolic pathways can ...be engineered more readily in cyanobacteria compared to higher photosynthetic organisms. Although significant progress has been made in pathway engineering, intracellular accumulation of the product is a potential bottleneck in large-scale production. Likewise, substrate uptake is known to limit growth and product formation. These limitations can potentially be addressed by targeted and controlled expression of transporter proteins in the metabolically engineered strains. This review focuses on the transporters that have been explored in cyanobacteria. To highlight the progress on characterization and application of cyanobacterial transporters, we applied text analytics to extract relevant information from over 1000 publications. We have categorized the transporters based on their source, their function and the solute they transport. Further, the review provides insights into the potential of transporters in the metabolic engineering of cyanobacteria for improved product titer.
•Extracting literature on cyanobacterial transporters using text analytics•Some heterologous transporters tested in cyanobacteria for strain improvement•Many native cyanobacterial transporters useful for production remain unexplored•Carbon and nitrogen transporters are widely studied for growth enhancement•Advanced bioinformatics tools need to be used to mine cyanobacterial transporters
Cyanobacteria provide an interesting platform for biotechnological applications due to their efficient photoautotrophic growth, amenability to genetic engineering and the ability to grow on ...non-arable land. An ideal industrial strain of cyanobacteria would need to be fast growing and tolerant to high levels of temperature, light, carbon dioxide, salt and be naturally transformable. In this study, we report Synechococcus elongatus PCC 11801, a strain isolated from India that fulfills these requirements. The physiological and biochemical characteristics of PCC 11801 under carbon and light-limiting conditions were investigated. PCC 11801 shows a doubling time of 2.3 h, that is the fastest growth for any cyanobacteria reported so far under ambient CO
conditions. Genome sequence of PCC 11801 shows genome identity of ~83% with its closest neighbors Synechococcus elongatus PCC 7942 and Synechococcus elongatus UTEX 2973. The unique attributes of PCC 11801 genome are discussed in light of the physiological characteristics that are needed in an industrial strain. The genome of PCC 11801 shows several genes that do not have homologs in neighbor strains PCC 7942 and UTEX 2973, some of which may be responsible for adaptation to various abiotic stresses. The remarkably fast growth rate of PCC 11801 coupled with its robustness and ease of genetic transformation makes it an ideal candidate for the photosynthetic production of fuels and chemicals.
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