Pseudomonas putida as a functional chassis for industrial biocatalysis: From native biochemistry to trans-metabolism

Abstract

The itinerary followed by Pseudomonas putida from being a soil-dweller and plant colonizer bacterium to become a flexible and engineer-able platform for metabolic engineering stems from its natural lifestyle, which is adapted to harsh environmental conditions and all sorts of physicochemical stresses. Over the years, these properties have been capitalized biotechnologically owing to the expanding wealth of genetic tools designed for deep-editing the P. putida genome. A suite of dedicated vectors inspired in the core tenets of synthetic biology have enabled to suppress many of the naturally-occurring undesirable traits native to this species while enhancing its many appealing properties, and also to import catalytic activities and attributes from other biological systems. Much of the biotechnological interest on P. putida stems from the distinct architecture of its central carbon metabolism. The native biochemistry is naturally geared to generate reductive currency [i.e., NAD(P)H] that makes this bacterium a phenomenal host for redox-intensive reactions. In some cases, genetic editing of the indigenous biochemical network of P. putida (cis-metabolism) has sufficed to obtain target compounds of industrial interest. Yet, the main value and promise of this species (in particular, strain KT2440) resides not only in its capacity to host heterologous pathways from other microorganisms, but also altogether artificial routes (trans-metabolism) for making complex, new-to-Nature molecules. A number of examples are presented for substantiating the worth of P. putida as one of the favorite workhorses for sustainable manufacturing of fine and bulk chemicals in the current times of the 4th Industrial Revolution. The potential of P. putida to extend its rich native biochemistry beyond existing boundaries is discussed and research bottlenecks to this end are also identified. These aspects include not just the innovative genetic design of new strains but also the incorporation of novel chemical elements into the extant biochemistry, as well as genomic stability and scaling-up issues.

Constitutively solvent‐tolerant Pseudomonas taiwanensis VLB120∆C∆ttgV supports particularly high‐styrene epoxidation activities when grown under glucose excess conditions

“…The central carbon metabolism of pseudomonads is naturally geared to generate plenty of reductive power, which makes these bacteria highly promising hosts for redox‐intensive reactions (Nikel & de Lorenzo, ). This is confirmed by the high specific epoxidation activities obtained with P. taiwanensis VLB120∆ C ∆ ttgV strains, which are unrivaled so far for NAD(P)H‐dependent microbial oxygenation under process conditions (Blank, Ebert, Buehler, & Bühler, ; Schrewe et al, ; Volmer et al, ).…”

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

“…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 ., ). 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 (Nikel and de Lorenzo, ). A collection of mutant strains obtained by random integration of mini‐Tn 5 elements, known as the Pseudomonas Reference Culture Collection, is available for strain KT2440 (Duque et al ., ).…”

“…The (somewhat short) list of bacterial hosts falling into this category includes Escherichia coli (Pontrelli et al ., ), Bacillus subtilis (Gu et al ., ), Streptomyces sp. (Spasic et al ., ), Pseudomonas putida (Nikel and de Lorenzo, ), and Corynebacterium glutamicum (Wendisch et al ., ), for which extensive background fundamental knowledge has been amassed. The wide use of these well‐established chassis notwithstanding, there has been a renewed interest for bringing up‐and‐coming host alternatives to the metabolic engineering community, thus broadening the repertoire of chassis available.…”

“…Tackling this problem is therefore one of the major challenges in bringing bacterial chassis to actual industrial‐scale bioproduction. In connection with this aspect, the evolution of alternative metabolic architectures in the engineered biochemical network (as opposed to changes operated merely at the genetic level) during bioproduction will help designing more efficient bioprocesses (Nikel and de Lorenzo, ).…”

Next‐Generation Industrial Biotechnology‐Transforming the Current Industrial Biotechnology into Competitive Processes

“…Many industrial microorganisms have been engineered as excellent production strains for CIB, such as Bacillus subtilis , E. coli , Corynebacterium glutamicum , Pseudomonas putida , Saccharomyces cerevisiae ( S. cerevisiae ) and so forth . Food supplements and fine chemicals such as vitamins, steroids, amino acids; biofuels including ethanol, butanol, biodiesel, and hydrogen; materials such as polyhydroxyalkanoates (PHA) and polysaccharides; polymer precursors like lactic acid (LA) succinic acid, and so forth.…”