Host Organism:<i>Pseudomonas putida</i>

Poblete‐Castro, Ignacio; Acuña, José Manuel Borrero‐de; Nikel, Pablo I.; Kohlstedt, Michael; Wittmann, Christoph

doi:10.1002/9783527807796.ch8

Cited by 25 publications

(17 citation statements)

References 180 publications

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“…The soil bacterium and platform strain Pseudomonas putida KT2440 has become the host of choice for metabolic engineering applications that require high levels of stress resistance and a robust, versatile metabolism (Nikel et al ., ; Poblete‐Castro et al ., ; Nikel and de Lorenzo, ). A dedicated synthetic biology toolbox is key to harness the full potential of this bacterium ― and strategies for tightly controlling gene expression are not only fundamental for understanding basic properties of the cell physiology and regulatory processes thereof, but they are also crucial for biotechnological applications (Gomez et al ., ; Benedetti et al ., ; Chavarría et al ., ; Martínez‐García and de Lorenzo, ; Calero and Nikel, ).…”

Section: Introductionmentioning

confidence: 99%

Physical decoupling of XylS/Pm regulatory elements and conditional proteolysis enable precise control of gene expression in Pseudomonas putida

Volke

Turlin

Mol

et al. 2019

Microbial Biotechnology

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Summary Most of the gene expression systems available for Gram‐negative bacteria are afflicted by relatively high levels of basal (i.e. leaky) expression of the target gene(s). This occurrence affects the system dynamics, ultimately reducing the output and productivity of engineered pathways and synthetic circuits. In order to circumvent this problem, we have designed a novel expression system based on the well‐known XylS/Pm transcriptional regulator/promoter pair from the soil bacterium Pseudomonas putida mt‐2, in which the key functional elements are physically decoupled. By integrating the xylS gene into the chromosome of the platform strain KT2440, while placing the Pm promoter into a set of standard plasmid vectors, the inducibility of the system (i.e. the output difference between the induced and uninduced state) improved up to 170‐fold. We further combined this modular system with an extra layer of post‐translational control by means of conditional proteolysis. In this setup, the target gene is tagged with a synthetic motif dictating protein degradation. When the system features were characterized using the monomeric superfolder GFP as a model protein, the basal levels of fluorescence were brought down to zero (i.e. below the limit of detection). In all, these novel expression systems constitute an alternative tool to altogether suppress leaky gene expression, and they can be easily adapted to other vector formats and plugged‐in into different Gram‐negative bacterial species at the user's will.

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Section: Introductionmentioning

confidence: 99%

Physical decoupling of XylS/Pm regulatory elements and conditional proteolysis enable precise control of gene expression in Pseudomonas putida

Volke

Turlin

Mol

et al. 2019

Microbial Biotechnology

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“…The advantages of this nonpathogenic soil bacterium for handling xeno-elements have been recently reviewed. [235] The well-known capacity of P. putida of growing in harsh environments (including industrial wastes) [236] and its tolerance to many organic solvents are traits that can be exploited to overcome the toxicity of compounds that can be exploited from neometabolism (e. g., fluoroacetate). Moreover, efficient mechanisms for organohalide dehalogenation [237] have been identified in this species and phylogenetically related isolates, suggesting that the biochemistry of P. putida has been evolutionarily shaped to deal with halogenated structures.…”

Section: Discussionmentioning

confidence: 99%

Intersecting Xenobiology and Neometabolism To Bring Novel Chemistries to Life

Nieto-Domínguez

Nikel

2020

ChemBioChem

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The diversity of life relies on a handful of chemical elements (carbon, oxygen, hydrogen, nitrogen, sulfur and phosphorus) as part of essential building blocks; some other atoms are needed to a lesser extent, but most of the remaining elements are excluded from biology. This circumstance limits the scope of biochemical reactions in extant metabolism – yet it offers a phenomenal playground for synthetic biology. Xenobiology aims to bring novel bricks to life that could be exploited for (xeno)metabolite synthesis. In particular, the assembly of novel pathways engineered to handle nonbiological elements (neometabolism) will broaden chemical space beyond the reach of natural evolution. In this review, xeno‐elements that could be blended into nature's biosynthetic portfolio are discussed together with their physicochemical properties and tools and strategies to incorporate them into biochemistry. We argue that current bioproduction methods can be revolutionized by bridging xenobiology and neometabolism for the synthesis of new‐to‐nature molecules, such as organohalides.

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“…The ubiquitous saprophytic, soil-colonizer P. putida, a Gram-negative, rod-shaped bacterium is increasingly being used as a chassis for applications characterized by harsh operating conditions. P. putida KT2440, which has been certified as a GRAS platform for recombinant protein production, is the most studied and used strain within the genus, and it is considered a safe host for cloning and expressing heterologous genes (Poblete-Castro et al, 2017). P. putida KT2440 possesses many of the desired features in an ideal bacterial chassis, such as rapid growth, low nutritional requirements and availability of sophisticated tools for genome and genetic manipulation .…”

Section: Pseudomonas Putidamentioning

confidence: 99%

Chasing bacterial chassis for metabolic engineering: a perspective review from classical to non‐traditional microorganisms

Calero

Nikel

2018

Microbial Biotechnology

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184

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The last few years have witnessed an unprecedented increase in the number of novel bacterial species that hold potential to be used for metabolic engineering. Historically, however, only a handful of bacteria have attained the acceptance and widespread use that are needed to fulfil the needs of industrial bioproduction - and only for the synthesis of very few, structurally simple compounds. One of the reasons for this unfortunate circumstance has been the dearth of tools for targeted genome engineering of bacterial chassis, and, nowadays, synthetic biology is significantly helping to bridge such knowledge gap. Against this background, in this review, we discuss the state of the art in the rational design and construction of robust bacterial chassis for metabolic engineering, presenting key examples of bacterial species that have secured a place in industrial bioproduction. The emergence of novel bacterial chassis is also considered at the light of the unique properties of their physiology and metabolism, and the practical applications in which they are expected to outperform other microbial platforms. Emerging opportunities, essential strategies to enable successful development of industrial phenotypes, and major challenges in the field of bacterial chassis development are also discussed, outlining the solutions that contemporary synthetic biology-guided metabolic engineering offers to tackle these issues.

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Host Organism:Pseudomonas putida

Cited by 25 publications

References 180 publications

Physical decoupling of XylS/Pm regulatory elements and conditional proteolysis enable precise control of gene expression in Pseudomonas putida

Physical decoupling of XylS/Pm regulatory elements and conditional proteolysis enable precise control of gene expression in Pseudomonas putida

Intersecting Xenobiology and Neometabolism To Bring Novel Chemistries to Life

Chasing bacterial chassis for metabolic engineering: a perspective review from classical to non‐traditional microorganisms

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