Synthesis of cefminox by cell-free extracts of<i>Streptomyces clavuligerus</i>

Kim, Jeong Keun; Kang, Heui Il; Chae, Jeong Seok; Park, Young Hoon; Choi, Yong Jin

doi:10.1111/j.1574-6968.2000.tb08914.x

Cited by 7 publications

(3 citation statements)

References 13 publications

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“…The cell-free rebooting of coliphage MS2, ΦX174, T7, and T4 using a phage genome has been reported ( 28 – 30 ). Methods for preparing a TXTL system from bacteria such as E. coli, Streptomyces spp ., P. putida, and Vibrio natriegens have also been reported ( 28 , 31 – 35 ) but are inadequate to accommodate cell-free creation of other phages. Although E. coli shows relatively high transformation efficiency, as a POC, we demonstrated the cell-free rebooting of a designer phage by using a TXTL system.…”

Section: Discussionmentioning

confidence: 99%

Synthetic engineering and biological containment of bacteriophages

Mitsunaka

Yamazaki

Pramono

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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The serious threats posed by drug-resistant bacterial infections and recent developments in synthetic biology have fueled a growing interest in genetically engineered phages with therapeutic potential. To date, many investigations on engineered phages have been limited to proof of concept or fundamental studies using phages with relatively small genomes or commercially available “phage display kits”. Moreover, safeguards supporting efficient translation for practical use have not been implemented. Here, we developed a cell-free phage engineering and rebooting platform. We successfully assembled natural, designer, and chemically synthesized genomes and rebooted functional phages infecting gram-negative bacteria and acid-fast mycobacteria. Furthermore, we demonstrated the creation of biologically contained phages for the treatment of bacterial infections. These synthetic biocontained phages exhibited similar properties to those of a parent phage against lethal sepsis in vivo. This efficient, flexible, and rational approach will serve to accelerate phage biology studies and can be used for many practical applications, including phage therapy.

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Section: Discussionmentioning

confidence: 99%

Synthetic engineering and biological containment of bacteriophages

Mitsunaka

Yamazaki

Pramono

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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“…With the shortest doubling time of all known organisms, Vibrio natriegens has garnered considerable interest as a promising microbial host to accelerate research and biotechnology. − Its rapid growth rate, which has been linked to high rates of protein synthesis and metabolic efficiency , suggests that this host may be harnessed as a powerful cell-free expression system. Cell-free bioproduction of protein and chemicals has been extensively investigated in E. coli and recently expanded to several other bacteria. − Accordingly, we assessed V. natriegens crude cell extract productivity in small-scale batch reactions using superfolder GFP (sfGFP) controlled by a T7 promoter. We systematically explored culturing conditions and extract additives to provide a simple and robust V. natriegens cell-free system.…”

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confidence: 99%

Establishing a Cell-Free Vibrio natriegens Expression System

et al. 2018

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The fast growing bacterium Vibrio natriegens is an emerging microbial host for biotechnology. Harnessing its productive cellular components may offer a compelling platform for rapid protein production and prototyping of metabolic pathways or genetic circuits. Here, we report the development of a V. natriegens cell-free expression system. We devised a simplified crude extract preparation protocol and achieved >260 μg/mL of superfolder GFP in a small-scale batch reaction after 3 h. Culturing conditions, including growth media and cell density, significantly affect translation kinetics and protein yield of extracts. We observed maximal protein yield at incubation temperatures of 26 or 30 °C, and show improved yield by tuning ions crucial for ribosomal stability. This work establishes an initial V. natriegens cell-free expression system, enables probing of V. natriegens biology, and will serve as a platform to accelerate metabolic engineering and synthetic biology applications.

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“…Together, these features have made cell-free systems an attractive platform for high-yield biomanufacturing of target compounds as well as for prototyping novel biosynthetic routes to their production. To date, several metabolic pathways and enzyme cascades have been implemented in cell-free formats including, but not limited to, those synthesizing antibiotics (Kim et al, 2000 ), cannabinoid precursors (Valliere et al, 2019 ), commodity alcohols (Guterl et al, 2012 ; Kay and Jewett, 2015 , 2020 ), food-grade antimicrobials (Kawai et al, 2003 ), glycolysis intermediates (Bogorad et al, 2013 ), hydrogen gas fuel (Martin Del Campo et al, 2013 ), isoprene compounds (Korman et al, 2014 ; Dudley et al, 2016 ), natural products (Goering et al, 2017 ), and nucleotides (Schultheisz et al, 2008 , 2011 ).…”

Section: Cell-free Biosynthesis Of Glycosylated Small Moleculesmentioning

confidence: 99%

Cell-Free Synthetic Glycobiology: Designing and Engineering Glycomolecules Outside of Living Cells

Jaroentomeechai

Taw

et al. 2020

Front. Chem.

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Glycans and glycosylated biomolecules are directly involved in almost every biological process as well as the etiology of most major diseases. Hence, glycoscience knowledge is essential to efforts aimed at addressing fundamental challenges in understanding and improving human health, protecting the environment and enhancing energy security, and developing renewable and sustainable resources that can serve as the source of next-generation materials. While much progress has been made, there remains an urgent need for new tools that can overexpress structurally uniform glycans and glycoconjugates in the quantities needed for characterization and that can be used to mechanistically dissect the enzymatic reactions and multi-enzyme assembly lines that promote their construction. To address this technology gap, cell-free synthetic glycobiology has emerged as a simplified and highly modular framework to investigate, prototype, and engineer pathways for glycan biosynthesis and biomolecule glycosylation outside the confines of living cells. From nucleotide sugars to complex glycoproteins, we summarize here recent efforts that harness the power of cell-free approaches to design, build, test, and utilize glyco-enzyme reaction networks that produce desired glycomolecules in a predictable and controllable manner. We also highlight novel cell-free methods for shedding light on poorly understood aspects of diverse glycosylation processes and engineering these processes toward desired outcomes. Taken together, cell-free synthetic glycobiology represents a promising set of tools and techniques for accelerating basic glycoscience research (e.g., deciphering the “glycan code”) and its application (e.g., biomanufacturing high-value glycomolecules on demand).

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Synthesis of cefminox by cell-free extracts ofStreptomyces clavuligerus

Cited by 7 publications

References 13 publications

Synthetic engineering and biological containment of bacteriophages

Synthetic engineering and biological containment of bacteriophages

Establishing a Cell-Free Vibrio natriegens Expression System

Cell-Free Synthetic Glycobiology: Designing and Engineering Glycomolecules Outside of Living Cells

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