Some photosynthetic organisms are capable of biosynthesizing carotenoids (xanthophylls) with α-carotene backbone, that is, α-carotene-derived carotenoids, such as (3R,3′R,6′R)-3,3′-dihydroxy α-carotene (lutein). Except for lutein, such carotenoids are minor compounds in nature. In this study, α-carotene-derived carotenoids were produced with E. coli. To achieve this, carotenoid biosynthesis genes from the bacterium Pantoea ananatis containing the 4-β-ketolase (crtW) gene with/without the 3-β-hydroxylase (crtZ) gene, in addition to crtEBI genes, and biosynthesis genes (MpLCYb, MpLCYe, and MpCYP97C) from liverwort Marchantia polymorpha, along with the HpIDI gene, were cloned into plasmids. The transformed E. coli cells biosynthesized (3S,3′R,6′R)-3,3′-dihydroxy-4-keto-α-carotene (fritschiellaxanthin (4-ketolutein)), (3′R,6′R)-3′-hydroxy-4-keto-α-carotene (4-keto-α-cryptoxanthin), and (3′R,6′R)-3′-hydroxy-α-carotene (α-cryptoxanthin), as carotenoids that have not been produced by a heterologous microbial system so far. These carotenoids show potent singlet oxygen-quenching activity.
Microorganisms have been extensively studied for their production of valuable chemicals. However, conventional gene fusion approaches often lack versatility and can result in enzyme inactivation. This study explored an alternative strategy for inducing metabolic channeling through sortase A-mediated ligation of metabolic enzymes. Sortase A recognizes specific amino acid sequences and selectively conjugates proteins at these sites. We focused on mevalonate production as a proof-of-concept to enhance the yield by assembling metabolic enzymes on a protein scaffold using sortase A. Although metabolic enzyme complexes were successfully formed using streptavidin as a scaffold, production did not improve. The use of CutA as a scaffold led to a 1.32-fold increase in production compared with that of the strain without the scaffold, demonstrating the efficacy of CutA in mevalonate production. These findings suggest that using sortase A to assemble metabolic enzymes onto a scaffold can effectively enhance microbial bioproduction.
Aldehydes are a class of compounds that contain carbonyl groups in their side chains and are widely used in industries such as fragrances, flavoring compounds, and pharmaceutical intermediates. In recent years, there has been a substantial rise in the application of microbial synthesis to generate aldehyde compounds and their derivatives. This review will conduct an in-depth analysis of the literature related to the manipulation of microorganisms for aldehyde accumulation and the subsequent generation of aldehyde-derived products using metabolic engineering and synthetic biology principles. Furthermore, the review further highlights the prospects and future potential of employing these aldehyde-accumulating microorganisms to synthesize a diverse range of value-added chemicals.
Synthetic biology, an emerging field at the intersection of biotechnology and engineering, has seen a global surge in application and awareness, necessitating a comprehensive understanding of its safe potentials to drive the bio-economy. This study aimed to assess the awareness and perceptions of synthetic biology among Nigerian biosciences stakeholders, including researchers, academicians, policymakers and students. The study employed a purposive online survey targeting diverse bioscience individuals and groups across Nigeria’s six geopolitical zones. The study received 107 responses from balanced gender representation with majority within the age group of 31–45 years old. The findings revealed a significant knowledge gap, with only 27.1% of respondents familiar with synthetic biology and 23.4% entirely unaware of it. Most respondents associated synthetic biology with biotechnology or genetic engineering and identified its applications to be in agriculture, medicine, environmental sustainability and research. Despite recognizing its benefits, many expressed concerns about safety, ethics, and regulation; notably, 43.9% of the respondents had concerns about synthetic biology with primary focus on safety and ethical implications. As regards the regulation of synthetic biology, the study showed that 80.4% of the respondents supported the need for synthetic biology regulation with few of the respondents (16.8%) aware of existing agency mandated to regulate synthetic biology. The respondents provided valuable insights into the various ways synthetic biology can be advanced in Nigeria which include increased awareness and capacity building, engagement through social media platforms, integration into education curricula and increased funding and investment in the research. The overall sentiment towards synthetic biology was positive, with 81.3% supporting its practice and 76.6% recognizing its positive global impact. However, a significant portion of respondents remained undecided. This study concludes that there is substantial gap in the knowledge of synthetic biology among bioscience stakeholders in Nigeria and the need for a heightened advocacy including continuous conferences and symposiums for the Nigeria bioscience community on the global potentials, concerns and regulation of synthetic biology. This will foster the acceptance of safe and responsible synthetic biology in Nigeria, thereby contributing to the broader national bio-economy development.
In most cyanobacteria, genetic engineering efforts currently rely upon chromosomal integration; a time-consuming process due to their polyploid nature. To enhance strain construction, here we develop and characterize two novel replicating plasmids for use in Synechococcus sp. PCC 7002. Following an initial screen of plasmids comprising seven different origins of replication, two were found capable of replication: one based on the WVO1 broad host range plasmid and the other a shuttle vector derived from pCB2.4 from Synechocystis sp. PCC 6803. These were then used to construct a set of new replicating plasmids, which were shown to be both co-transformable and stably maintained in PCC 7002 at copy numbers between 7–16 and 0.6–1.4, respectively. Lastly, we demonstrate the importance of using multimeric plasmids during natural transformation of PCC 7002, with higher order multimers providing a 30-fold increase in transformation efficiency relative to monomeric plasmids. Useful considerations and methods for enhancing multimer content in plasmid samples are also presented.
The desire to harness nature’s capability of precise gene expression regulation has motivated the pursuit of synthetic gene circuits. However, designing and building novel synthetic gene circuits with predictable dynamics is nontrivial. To facilitate the design, cell-free systems have emerged as an effective alternative testbed to living biological systems in characterizing and prototyping synthetic gene regulatory networks, given its relative simplicity and designability in terms of cellular contents. Meanwhile, as parameterizing and analyzing first principle-based models can shed light on the required kinetic parameter values, thus the specific regulatory components, for the desired dynamics, coupling mathematical modeling with cell-free experiments has become an effective approach in exploring novel synthetic gene circuits. In this mini-review, we provide an overview of current progress on using deterministic first principle-based mathematical modeling in conjunction with cell-free systems, in designing and characterizing novel gene circuits, as well as the standing challenges and issues with this approach.
Polyketides (PKs) are a large class of secondary metabolites produced by microorganisms and plants, characterized by highly diverse structures and broad biological activities. They have wide market and application prospects in medicine, agriculture, and the food industry. The complex chemical structures and multiple steps of natural polyketides result in yield that cannot be met by purely synthetic methods. With the development of synthetic biology, a number of novel technologies and synthetic strategies have been developed for the efficient synthesis of polyketides. This paper first introduces polyketides from different sources and classifications, then the reconstruction of biosynthetic pathways is described using a “bottom-up” synthetic biology approach. Through methods such as enhancing precursors, relieving feedback inhibition, and dynamic regulation, the efficient production of polyketides is achieved. Finally, the challenges faced by polyketides research and future development directions are discussed.
Cupriavidus necator H16 has been intensively explored for its potential as a versatile microbial cell factory, especially for its CO2 fixation capability over the past few decades. However, rational metabolic engineering remains challenging in the construction of microbial cell factories with complex phenotypes due to the limited understanding of its metabolic regulatory network. To overcome this obstacle, laboratory adaptive evolution emerges as an alternative. In the present study, CAM (cytosine deaminase-assisted mutator) was established for the genome evolution of C. necator, addressing the issue of low mutation rates. By fusing cytosine deaminase with single-stranded binding proteins, CAM introduced genome-wide C-to-T mutations during DNA replication. This innovative approach could boost mutation rates, thereby expediting laboratory adaptive evolution. The applications of CAM were demonstrated in improving cell factory robustness and substrate utilization, with H2O2 resistance and ethylene glycol utilization as illustrative case studies. This genetic tool not only facilitates the development of efficient cell factories but also opens avenues for exploring the intricate phenotype-genotype relationships in C. necator.
As an important
intermediate in the tricarboxylic acid (TCA) cycle,
Advancements in the Bio-degradation of Plastic Waste into Value-added Chemicals: A Recent Perspective
Plastics are an essential component of modern life, but the plastic waste has caused significant environmental pollution and economic losses. The effective solution to these problems is the biodegradation and high-value conversion of plastic waste. After biodegradation, plastic waste is broken into smaller molecules and eventually transformed into innocuous substances like water, carbon dioxide and biomass. High-value conversion enables plastic waste to be converted into products with higher economic value and environmental friendliness. Based on this, we summarize the biodegradation methods of bioplastics and analyze the shortage of these methods. Subsequently, we summarize the progress of converting the degradation products into value-added chemicals, comprehensively analyze the advantages and disadvantages of these bioconversion process, and propose some strategies to address these disadvantages. Finally, we analyze the significance of establishing a microbial-based conversion process that integrates the degradation and the conversion, and propose some potential strategies.
