scholarly journals Biotechnological Production of the Sunscreen Pigment Scytonemin in Cyanobacteria: Progress and Strategy

Marine Drugs ◽  
2021 ◽  
Vol 19 (3) ◽  
pp. 129
Author(s):  
Xiang Gao ◽  
Xin Jing ◽  
Xufeng Liu ◽  
Peter Lindblad
Keyword(s):  
High Efficiency ◽  
Cost Effective ◽  
Gene Clusters ◽  
Central Carbon Metabolism ◽  
Biochemical Properties ◽  
High Yield ◽  
Nostoc Punctiforme ◽  
Biosynthetic Gene Clusters ◽  
Acid Biosynthesis ◽  
Biotechnological Production

Scytonemin is a promising UV-screen and antioxidant small molecule with commercial value in cosmetics and medicine. It is solely biosynthesized in some cyanobacteria. Recently, its biosynthesis mechanism has been elucidated in the model cyanobacterium Nostoc punctiforme PCC 73102. The direct precursors for scytonemin biosynthesis are tryptophan and p-hydroxyphenylpyruvate, which are generated through the shikimate and aromatic amino acid biosynthesis pathway. More upstream substrates are the central carbon metabolism intermediates phosphoenolpyruvate and erythrose-4-phosphate. Thus, it is a long route to synthesize scytonemin from the fixed atmospheric CO2 in cyanobacteria. Metabolic engineering has risen as an important biotechnological means for achieving sustainable high-efficiency and high-yield target metabolites. In this review, we summarized the biochemical properties of this molecule, its biosynthetic gene clusters and transcriptional regulations, the associated carbon flux-driving progresses, and the host selection and biosynthetic strategies, with the aim to expand our understanding on engineering suitable cyanobacteria for cost-effective production of scytonemin in future practices.

scholarly journals Salicylic Acid Biosynthesis and Metabolism: A Divergent Pathway for Plants and Bacteria

Biomolecules ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 705
Author(s):  
Awdhesh Kumar Mishra ◽  
Kwang-Hyun Baek
Keyword(s):  
Salicylic Acid ◽  
Bacterial Species ◽  
Shikimate Pathway ◽  
Gene Clusters ◽  
Defense Responses ◽  
Biosynthetic Gene ◽  
Biosynthetic Enzyme ◽  
Biosynthetic Gene Clusters ◽  
Acid Biosynthesis ◽  
Common Precursor

Salicylic acid (SA) is an active secondary metabolite that occurs in bacteria, fungi, and plants. SA and its derivatives (collectively called salicylates) are synthesized from chorismate (derived from shikimate pathway). SA is considered an important phytohormone that regulates various aspects of plant growth, environmental stress, and defense responses against pathogens. Besides plants, a large number of bacterial species, such as Pseudomonas, Bacillus, Azospirillum, Salmonella, Achromobacter, Vibrio, Yersinia, and Mycobacteria, have been reported to synthesize salicylates through the NRPS/PKS biosynthetic gene clusters. This bacterial salicylate production is often linked to the biosynthesis of small ferric-ion-chelating molecules, salicyl-derived siderophores (known as catecholate) under iron-limited conditions. Although bacteria possess entirely different biosynthetic pathways from plants, they share one common biosynthetic enzyme, isochorismate synthase, which converts chorismate to isochorismate, a common precursor for synthesizing SA. Additionally, SA in plants and bacteria can undergo several modifications to carry out their specific functions. In this review, we will systematically focus on the plant and bacterial salicylate biosynthesis and its metabolism.

scholarly journals Recent Advances in Strategies for the Cloning of Natural Product Biosynthetic Gene Clusters

2021 ◽  
Vol 9 ◽  
Author(s):  
Wenfang Wang ◽  
Guosong Zheng ◽  
Yinhua Lu
Keyword(s):  
High Efficiency ◽  
Therapeutic Potential ◽  
Gene Clusters ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
High Molecular Weight Dna ◽  
Discovery Research ◽  
Recent Advances ◽  
Drug Discovery Research ◽  
Dna Fragment

Microbial natural products (NPs) are a major source of pharmacological agents. Most NPs are synthesized from specific biosynthetic gene clusters (BGCs). With the rapid increase of sequenced microbial genomes, large numbers of NP BGCs have been discovered, regarded as a treasure trove of novel bioactive compounds. However, many NP BGCs are silent in native hosts under laboratory conditions. In order to explore their therapeutic potential, a main route is to activate these silent NP BGCs in heterologous hosts. To this end, the first step is to accurately and efficiently capture these BGCs. In the past decades, a large number of effective technologies for cloning NP BGCs have been established, which has greatly promoted drug discovery research. Herein, we describe recent advances in strategies for BGC cloning, with a focus on the preparation of high-molecular-weight DNA fragment, selection and optimization of vectors used for carrying large-size DNA, and methods for assembling targeted DNA fragment and appropriate vector. The future direction into novel, universal, and high-efficiency methods for cloning NP BGCs is also prospected.

scholarly journals A Streptomyces venezuelae Cell-Free Toolkit for Synthetic Biology

2020 ◽  
Author(s):  
Simon J Moore ◽  
Hung-En Lai ◽  
Soo-Mei Chee ◽  
Ming Toh ◽  
Seth Coode ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Gene Clusters ◽  
Test Tube ◽  
High Yield ◽  
Proof Of Concept ◽  
Biosynthetic Gene Clusters ◽  
Reaction Conditions ◽  
One Pot ◽  
Free System ◽  
High Level

AbstractProkaryotic cell-free coupled transcription-translation (TX-TL) systems are emerging as a powerful tool to examine natural product biosynthetic pathways in a test-tube. The key advantages of this approach are the reduced experimental timescales and controlled reaction conditions. In order to realise this potential, specialised cell-free systems in organisms enriched for biosynthetic gene clusters, with strong protein production and well-characterised synthetic biology tools, is essential. The Streptomyces genus is a major source of natural products. To study enzymes and pathways from Streptomyces, we originally developed a homologous Streptomyces cell-free system to provide a native protein folding environment, a high G+C (%) tRNA pool and an active background metabolism. However, our initial yields were low (36 μg/mL) and showed a high level of batch-to-batch variation. Here, we present an updated high-yield and robust Streptomyces TX-TL protocol, reaching up to yields of 266 μg/mL of expressed recombinant protein. To complement this, we rapidly characterise a range of DNA parts with different reporters, express high G+C (%) biosynthetic genes and demonstrate an initial proof of concept for combined transcription, translation and biosynthesis of Streptomyces metabolic pathways in a single ‘one-pot’ reaction.

scholarly journals The Application of Ribosome Engineering to Natural Product Discovery and Yield Improvement in Streptomyces

Antibiotics ◽  
2019 ◽  
Vol 8 (3) ◽  
pp. 133 ◽  
Author(s):  
Saibin Zhu ◽  
Yanwen Duan ◽  
Yong Huang
Keyword(s):  
Natural Product ◽  
Ribosomal Proteins ◽  
Cost Effective ◽  
Gene Clusters ◽  
Product Formation ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Yield Improvement ◽  
Ribosome Engineering ◽  
Natural Product Discovery

Microbial natural product drug discovery and development has entered a new era, driven by microbial genomics and synthetic biology. Genome sequencing has revealed the vast potential to produce valuable secondary metabolites in bacteria and fungi. However, many of the biosynthetic gene clusters are silent under standard fermentation conditions. By rational screening for mutations in bacterial ribosomal proteins or RNA polymerases, ribosome engineering is a versatile approach to obtain mutants with improved titers for microbial product formation or new natural products through activating silent biosynthetic gene clusters. In this review, we discuss the mechanism of ribosome engineering and its application to natural product discovery and yield improvement in Streptomyces. Our analysis suggests that ribosome engineering is a rapid and cost-effective approach and could be adapted to speed up the discovery and development of natural product drug leads in the post-genomic era.

scholarly journals Characterization of Cas proteins for CRISPR-Cas editing in streptomycetes

2019 ◽  
Author(s):  
Wan Lin Yeo ◽  
Elena Heng ◽  
Lee Ling Tan ◽  
Yi Wee Lim ◽  
Yee Hwee Lim ◽  
...  
Keyword(s):  
Transcriptional Regulation ◽  
Genome Editing ◽  
High Efficiency ◽  
Streptococcus Thermophilus ◽  
Gene Clusters ◽  
Biosynthetic Gene Clusters ◽  
Tularensis Subsp ◽  
Homology Directed Repair ◽  
Cas Proteins

Application of the well-characterized Streptococcus pyogenes CRISPR-Cas9 system in actinomycetes has enabled high efficiency multiplex genome editing and CRISPRi-mediated transcriptional regulation in these prolific bioactive metabolite producers. Nonetheless, SpCas9 has its limitations and can be ineffective depending on the strains and target sites. Here, we built and tested alternative CRISPR-Cas constructs based on the standalone pCRISPomyces-2 editing plasmid. We showed that Streptococcus thermophilus CRISPR1 (sth1Cas9), Staphylococcus aureus Cas9 (saCas9), and Francisella tularensis subsp. Novicida U112 Cpf1 (fnCpf1) are functional in multiple streptomycetes enabling efficient homology directed repair (HDR)-mediated knock-in and deletion. In strains where spCas9 was non-functional, these alternative Cas systems enabled precise genomic modifications within biosynthetic gene clusters for the discovery, production and diversification of natural products. These additional Cas proteins provide us with the versatility to overcome the limitations of individual CRISPR-Cas systems for genome editing and transcriptional regulation of these industrially important bacteria.

scholarly journals High-Efficiency Production of Large-Size Few-Layer Graphene Platelets via Pulsed Discharge of Graphite Strips

Nanomaterials ◽  
2019 ◽  
Vol 9 (12) ◽  
pp. 1785 ◽  
Author(s):  
Xin Gao ◽  
Tomomasa Hiraoka ◽  
Shunsuke Ohmagari ◽  
Shigeru Tanaka ◽  
Zemin Sheng ◽  
...  
Keyword(s):  
High Efficiency ◽  
Expanded Graphite ◽  
Cost Effective ◽  
High Yield ◽  
Pulsed Discharge ◽  
Force Microscopy ◽  
Large Size ◽  
Potential Applications ◽  
Graphene Platelets ◽  
Few Layer Graphene

The synthesis of large-size graphene materials is still a central focus of research into additional potential applications in various areas. In this study, large-size graphene platelets were successfully produced by pulsed discharge of loose graphite strips with a dimension of 2 mm × 0.5 mm × 80 mm in distilled water. The graphite strips were made by pressing and cutting well-oriented expanded graphite paper. The recovered samples were characterized by various techniques, including TEM, SEM, optical microscopy (OM), atomic force microscopy (AFM), XRD and Raman spectroscopy. Highly crystalline graphene platelets with a lateral dimension of 100–200 μm were identified. The high yield of recovered graphene platelets is in a range of 90–95%. The results also indicate that increasing charging voltage improves the yield of graphene platelets and decreases the number of graphitic layers in produced graphene platelets. The formation mechanism of graphene platelets was discussed. This study provides a one-step cost-effective route to prepare highly crystalline graphene platelets with a sub-millimeter lateral size.

A Review: Klebsiella pneumoniae, Klebisellaoxytoca and Biotechnology

2021 ◽  
Vol 26 (3) ◽  
pp. 2567-2586
Author(s):  
CATERINA TOMULESCU ◽  
◽  
MIȘU MOSCOVICI ◽  
IRINA LUPESCU ◽  
ROXANA MĂDĂLINA STOICA ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Genetic Manipulation ◽  
Klebsiella Oxytoca ◽  
Cost Effective ◽  
Biochemical Properties ◽  
High Yield ◽  
Natural Environments ◽  
Klebsiella Species ◽  
Bulk Chemicals ◽  
Plant Growth Promoting Activities

Biotechnology, molecular biology and genetic engineering, and bioprospecting play a crucial role in our common future, enabling industrially important microorganisms to ensure sustainable products (fuels, chemicals, pharmaceuticals, food, drug delivery systems, medical devices etc.) and new bioeconomic opportunities. Biotechnological applications are able to provide cost-effective green alternatives to conventional industrial processes, which are currently affecting the nature and biodiversity. Klebsiella species are among the well-studied microbes both in medicine field, as ones of the most resilient opportunistic pathogens, and in industry, due to their promising biochemical properties, and their potential as better hosts than other microorganisms, for i.e. in genetic manipulation. Klebsiella oxytoca and Klebsiella pneumoniae are ubiquitously found in natural environments, but also as commensals in the human gut, and associated with a high-resistance to the first-line antibiotics. However, these specific strains are continuously isolated and studied for different industrial purposes (i.e. bulk chemicals and biofuels production, medical diagnosis, nanoparticles and exopolysaccharides synthesis, plant growth promoting activities, bioremediation and biodegradation agents etc.), and scientific results regarding their biotechnological potential could generate big impact for global bioeconomy development. Recently, research in synthetic biology gained a lot of attention, and new techniques highlight ways to reprogramme these microbial cells in view of high-yield or high-quality new chemicals obtainment. Therefore, some scientific research niches are emerging in biotechnology, and unknown metabolic pathways and genes are identified and further studied, to provide alternative solutions to the global challenges.

scholarly journals Expanding gene families helps generate the metabolic robustness required for antibiotic biosynthesis

2017 ◽  
Author(s):  
Jana K Schniete ◽  
Pablo Cruz-Morales ◽  
Nelly Selem ◽  
Lorena T. Fernández-Martínez ◽  
Iain S Hunter ◽  
...  
Keyword(s):  
Gene Families ◽  
Gene Clusters ◽  
Central Carbon Metabolism ◽  
Metabolic Adaptation ◽  
Antibiotic Biosynthesis ◽  
Biosynthetic Gene Clusters ◽  
Metabolic Plasticity ◽  
Central Carbon ◽  
Metabolic Robustness ◽  
Cell Functionality

Introductory paragraphExpanding the genetic repertoire of an organism by gene duplication or horizontal gene transfer (HGT) can aid adaptation. Streptomyces species are prolific producers of bioactive specialised metabolites with adaptive functions in nature and some have found utility in human medicine such as antibiotics. Whilst the biosynthesis of these specialised metabolites is directed by dedicated biosynthetic gene clusters (BGCs), little attention has been focussed on how these organisms have evolved robustness into their genomes to facilitate the metabolic plasticity required to provide chemical precursors for biosynthesis. Here we show that specific expansions of gene families in central carbon metabolism have evolved and become fixed in Streptomyces bacteria to enable plasticity and robustness that maintain cell functionality whilst costly specialised metabolites are produced. These expanded gene families, in addition to being a metabolic adaptation, make excellent targets for metabolic engineering of industrial specialised metabolite producing bacteria.

Expanding the Natural Products Heterologous Expression Repertoire in the Model Cyanobacterium Anabaena sp. Strain PCC 7120: Production of Pendolmycin and Teleocidin B-4

2019 ◽  
Author(s):  
Patrick Videau ◽  
Kaitlyn Wells ◽  
Arun Singh ◽  
Jessie Eiting ◽  
Philip Proteau ◽  
...  
Keyword(s):  
Natural Products ◽  
Genome Mining ◽  
Gene Clusters ◽  
Combinatorial Biosynthesis ◽  
Test Case ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Cyanobacterium Anabaena ◽  
Anabaena Sp ◽  
Pcc 7120

Cyanobacteria are prolific producers of natural products and genome mining has shown that many orphan biosynthetic gene clusters can be found in sequenced cyanobacterial genomes. New tools and methodologies are required to investigate these biosynthetic gene clusters and here we present the use of <i>Anabaena </i>sp. strain PCC 7120 as a host for combinatorial biosynthesis of natural products using the indolactam natural products (lyngbyatoxin A, pendolmycin, and teleocidin B-4) as a test case. We were able to successfully produce all three compounds using codon optimized genes from Actinobacteria. We also introduce a new plasmid backbone based on the native <i>Anabaena</i>7120 plasmid pCC7120ζ and show that production of teleocidin B-4 can be accomplished using a two-plasmid system, which can be introduced by co-conjugation.

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