Patchoulol Production with Metabolically Engineered Corynebacterium glutamicum

Abstract

Patchoulol is a sesquiterpene alcohol and an important natural product for the perfume industry. Corynebacterium glutamicum is the prominent host for the fermentative production of amino acids with an average annual production volume of ~6 million tons. Due to its robustness and well established large-scale fermentation, C. glutamicum has been engineered for the production of a number of value-added compounds including terpenoids. Both C40 and C50 carotenoids, including the industrially relevant astaxanthin, and short-chain terpenes such as the sesquiterpene valencene can be produced with this organism. In this study, systematic metabolic engineering enabled construction of a patchoulol producing C. glutamicum strain by applying the following strategies: (i) construction of a farnesyl pyrophosphate-producing platform strain by combining genomic deletions with heterologous expression of ispA from Escherichia coli; (ii) prevention of carotenoid-like byproduct formation; (iii) overproduction of limiting enzymes from the 2-c-methyl-d-erythritol 4-phosphate (MEP)-pathway to increase precursor supply; and (iv) heterologous expression of the plant patchoulol synthase gene PcPS from Pogostemon cablin. Additionally, a proof of principle liter-scale fermentation with a two-phase organic overlay-culture medium system for terpenoid capture was performed. To the best of our knowledge, the patchoulol titers demonstrated here are the highest reported to date with up to 60 mg L−1 and volumetric productivities of up to 18 mg L−1 d−1.

Construction of yeast producing patchoulol by global metabolic engineering strategy

“…Furthermore, using plants as a source for producing a sesquiterpene includes slow growth and varying compositions depending on the geographical location and climate conditions. Several studies have been reported using recombinant microbial cells to produce patchoulol with less time and energy consumption (Albertsen et al, 2011; Asadollahi et al, 2008; Henke et al, 2018; Ma et al, 2019). Asadollahi et al (2008) achieved the patchoulol production of 11.5 mg/L from galactose by suppressing ERG9 after introducing a patchoulol synthesis gene in S. cerevisiae .…”

Optimization of factors influencing enzyme activity and product selectivity and the role of proton transfer in the catalytic mechanism of patchoulol synthase

“…Patchouli is an aromatic plant with essential oils that is cultivated in the subtropical regions of India, China, and Southeast Asia like Indonesia. Patchouli oil has immense export potential due to its low production costs and high demand in the perfume and cosmetics industry . Usually the production is done in small companies where the equipment does not meet European standards and there are less regulatory oversights.…”

Impact of CRISPR interference on strain development in biotechnology

“…The model organism C. glutamicum is widely used in biotechnological industry, since it is a workhorse able to produce various compounds with high production titers [106–117], especially the amino acids l ‐glutamate [118–121] and l ‐lysine [122–127]. Globally, amino acids from C. glutamicum are produced in million ton scale [109].…”