Gram‐scale production of the sesquiterpene α‐humulene with Cupriavidus necator

Abstract

Terpenoids have an impressive structural diversity and provide valuable substances for a variety of industrial applications. Among terpenes, the sesquiterpenes (C 15 ) are the largest subclass with bioactivities ranging from aroma to health promotion. In this article, we show a gram-scale production of the sesquiterpene α-humulene in final aqueous concentrations of 2 g L −1 with the recombinant strain Cupriavidus necator pKR-hum in a fed-batch mode on fructose as carbon source and n-dodecane as an extracting organic phase for in situ product removal. Since C. necator is capable of both heterotrophic and autotrophic growth, we additionally modeled the theoretically possible yields of a heterotrophic versus an autotrophic process on CO 2 in industrially relevant quantities. We compared the cost-effectiveness of both processes based on a production of 10 t α-humulene per year, with both processes performing equally with similar costs and gains. Furthermore, the expression and activity of 3-hydroxymethylglutaryl-CoA reductase (hmgR) from Myxococcus xanthus was identified as the main limitation of our constructed C. necator pKR-hum strain.Thus, we outlined possible solutions for further improvement of our production strain, for example, the replacement of the hmgR from M. xanthus by a plant-based variant to increase α-humulene production titers in the future.

Effects of Cabya (Piper retrofractum Vahl.) Fruit Developmental Stage on VOCs

“…It is also currently considered suitable for the construction of natural product-like compounds through diversity-directed synthesis; using natural products as a starting point in this process is an emerging strategy for constructing diverse backbones to meet the need for high-throughput screening in drug development [ 32 ]. As part of the pepper scent, the sesquiterpenes, such as α-humulene and germacrene D, which make up the biggest subgroup of terpenes, also have biological properties that are beneficial to health [ 33 , 34 ]. α-humulene has specifically been shown to have anti-inflammatory and anti-cancer activities, with great potential for medical applications [ 35 ].…”

Synthetic biology toolkit for engineering Cupriviadus necator H16 as a platform for CO2 valorization

“…Sesquiterpenes (C15), the largest subgroup of terpenoids, has a wide application in industry, such as β-farnesene, a precursor for a jet fuel additive, and α-humulene, a potential drug to treat cancer. Milker et al obtained 26.3 ± 1.3 µM β-farnesene [ 173 ] and 2 g/L α-humulene [ 174 ] in a fed-batch mode on fructose as carbon source by expressing β-farnesene synthase and α-humulene synthase, respectively. Both studies showed that additional MVA expression contributed little to increase the titer of terpenoids, which might be resulted from the poor expression of hydroxymethylglutaryl-CoA reductase ( hmgR ).…”

Dual cytoplasmic‐peroxisomal engineering for high‐yield production of sesquiterpene α‐humulene in Yarrowia lipolytica

“…With the fast development of synthetic biology, the engineered microbial cell factory was gradually considered to be a promising strategy to produce a variety of high‐value compounds on an industrial scale, including α‐humulene. Previous studies mainly focused on engineered Escherichia coli , Saccharomyces cerevisiae , and Cupriavidus necator for heterologous α‐humulene production (Alemdar et al, 2017; Arnesen et al, 2020; Harada et al, 2009; Krieg et al, 2018; Milker et al, 2021; Sonntag et al, 2015; Zhang et al, 2018, Zhang et al, 2020). However, α‐humulene titer in these engineered microbes was still too low (~2 g/l).…”

“…necator for heterologous α-humulene production (Alemdar et al, 2017;Arnesen et al, 2020;Harada et al, 2009;Krieg et al, 2018;Milker et al, 2021;Sonntag et al, 2015;Zhang et al, 2018, Zhang et al, 2020. However, α-humulene titer in these engineered microbes was still too low (~2 g/l).…”