Metabolic engineering of Synechocystis sp. PCC 6803 for the photoproduction of the sesquiterpene valencene

Abstract

Cyanobacteria are extremely adaptable, fast-growing, solar-powered cell factories that, like plants, are able to convert carbon dioxide into sugar and oxygen and thereby produce a large number of important compounds. Due to their unique phototrophy-associated physiological properties, i.e. naturally occurring isoprenoid metabolic pathway, they represent a highly promising platform for terpenoid biosynthesis. Here, we implemented a carefully devised engineering strategy to boost the biosynthesis of commercially attractive plant sequiterpenes, in particular valencene. Sesquiterpenes are a diverse group of bioactive metabolites, mainly produced in higher plants, but with often low concentrations and expensive downstream extraction. In this work we successfully demonstrate a multi-component engineering approach towards the photosynthetic production of valencene in the cyanobacterium Synechocystis sp. PCC 6803. First, we improved the flux towards valencene by markerless genomic deletions of shc and sqs . Secondly, we downregulated the formation of carotenoids, which are essential for viability of the cell, using CRISPRi on crtE . Finally, we intended to increase the spatial proximity of the two enzymes, ispA and CnVS , involved in valencene formation by creating an operon construct, as well as a fusion protein. Combining the most successful strategies resulted in a valencene production of 19 mg/g DCW in Synechocystis . In this work, we have devised a useful platform for future engineering steps.

Probing the Synergistic Ratio of P450/CPR To Improve (+)-Nootkatone Production in Saccharomyces cerevisiae

“…There are numerous reports exploiting whole-cell biotransformation systems to achieve a high titer and productivity to achieve cost competitiveness (Table ). Compared with the reported results, ,,,, the highest volumetric titers of 539.3 and 208 mg/L for (+)-valencene and (+)-nootkatone were reported by Wriessnegger et al with productivities of 3.96 and 1.92 mg/Lh, respectively. In this study, the highest (+)-valencene and (+)-nootkatone titers were dramatically elevated to 3.73 and 1.02 g/L by the PK2RI-AtC/H m6 A strain, indicating that this is an economically viable route for it in the g/L range.…”

Green Synthesis of Silver Nanoparticles by the Cyanobacteria Synechocystis sp.: Characterization, Antimicrobial and Diabetic Wound-Healing Actions

“…The cyanobacterium Synechocystis sp. are rich in entities/bioactive compounds, which have reducing power for ions within their vicinity turning them into ecofriendly nanoparticles [27,28]. For instance, Synechocystis sp.…”

CRISPR-Based Approaches for Gene Regulation in Non-Model Bacteria

“…These tools can be used to tune endogenous metabolism and/or heterologous metabolic pathways ( Peng et al, 2018 ; Banerjee et al, 2020 ). CRISPRi/a tools are most often combined with other metabolic engineering techniques, such as the deletion, overexpression, or mutation of select genes and optimization of medium, to further increase titers of the desired product ( Park et al, 2019 ; Dietsch et al, 2021 ; Kozaeva et al, 2021 ).…”