ReviewRecent advances in microbial production of phenolic compounds
Introduction
Phenolic compounds (PCs) comprise a diverse group of plant secondary metabolites, and can be categorized into polyphenols, phenolic acids, phenylpropanoids, flavonoids, stilbenes, coumarins and their derivatives (e.g. esters, glycosides and polymers) [1]. PCs possess the aromatic ring(s) bearing one or more hydroxyl groups, and exhibit multifaceted activities (antioxidation, anti-inflammation, anti-tumor, antiviral, anti-bacteria and so on) [2,3]. Due to these health-beneficial properties, PCs are widely used in nutraceutical, pharmaceutical and cosmetic industries.
Currently, PCs are mainly manufactured by either plant extraction or chemical synthesis. However, the plant extraction process often encounters bottlenecks such as low yield, fluctuation of active ingredients caused by seasonal/climatic variations, and over-exploitation of plant resources, and thus is difficult to fulfill the growing demands. Chemical synthesis also faces problems such as expensive precursors, inadequate regio- and stereo-selectivity, use of toxic catalysts and harsh reaction conditions. Therefore, engineering microbial cell factories has become a promising alternative approach to produce these high-value PCs.
PCs like other aromatic compounds are generally biosynthesized through the shikimate pathway. This pathway consists of seven enzymatic steps that convert the precursors erythrose-4-phosphate (E4P) and phosphoenolpyruvate (PEP) to chorismate. The involved enzymes and the underlying regulation mechanism are well elucidated, and numerous studies have been conducted to boost the shikimate pathway for the production of various aromatic compounds by metabolic strategies like increasing the supply of PEP and E4P, alleviating feedback inhibition of end-products on upstream enzymes, and blocking/weakening the competing pathways. These strategies are well summarized already [[4], [5], [6]] and thus will not be the emphasis of this review.
The rapid expansion of available information on plant genomes and transcriptomes has accelerated enzyme mining and identification of the biosynthetic pathways from the native species. However, so far there are many PCs whose native pathways are still elusive. Fortunately, artificial pathways can be designed by mining genes from different species based on the bioinformatics databases. To meet the industrial application requirements, the titer, yield and productivity of the recombinant microorganisms need to be improved. To this end, the advancements in protein engineering, metabolic engineering and synthetic biology provide powerful tools to remove obstacles in the construction of microbial cell factories to produce PCs. In this review, we summarize the recent progress on the design of artificial pathways for the biosynthesis of PCs, and the strategies used to improve the production efficiency, which include enzyme mining and engineering, modular pathway assembly and optimization, coculture engineering, and dynamic regulation of carbon flux (Table 1).
Section snippets
Enzyme Mining to Construct Artificial Pathways
To achieve heterologous production of a PC, a straightforward way is to reconstruct the entire native pathway in a genetic tractable microorganism. However, in many cases the native pathway may be partially characterized or even completely unknown. Thanks to the enormous enzyme information documented in the databases as BRENDA, one can dig up alternative enzymes from other organisms to fill the gap(s) in a pathway, or even design artificial pathways by connecting the target product with the
Balancing Enzyme Expression to Improve Pathway Efficiency
Flavonoids comprise a highly diverse group of plant secondary metabolites, which can be further categorized into flavones, flavonols, flavonones, isoflavones, anthocyanins and catechins according to their structure variability. Due to their diverse biological properties, flavonoids have been widely used in human health and nutrition [31]. To achieve economical and sustainable production of flavonoids, increasing efforts have been made to improve the efficiency of the biosynthetic pathways.
Coculture Engineering to Alleviate Metabolic Burden and Unwanted Side-reactions
Modular optimization proved an effective strategy to balance the biosynthetic pathways. However, introducing long pathways in a single strain may cause severe metabolic burden. Thus, coculture engineering has been used to improve the biosynthesis of a variety of PCs. Compared with monocultures, cocultures shows several advantages, including division of labor to alleviate cell burden, compartmentalization of reactions to reduce metabolic interference [33].
Salidroside, a glucoside of tyrosol, is
Genetic Circuit for Dynamic Regulation and High Throughput Screening
The objective of microbial production is to maximize the titer, yield and productivity of the engineered strains, which requires systematic remodeling of cell metabolism to coordinate the distribution of metabolic flux between growth and production. Oftentimes, too early expression of pathway enzymes and inhibition of competing pathways can impair cell growth.
Toggle switches have been used to shift the cell status by adding small molecule inducers. Malonyl-CoA is a versatile building block for
Challenges and Perspectives
Although significant progress has been made in microbial production of PCs, many challenges still need to be overcome on the way to their economical production. First, many PCs are derived from plants, and some types of plant enzymes such as cytochrome P450s are often poorly expressed in yeast or bacteria. Thus, the sequence, codon usage, expression level and subcellular localization usually need to be optimized. Emerging new tools such as machine learning [52] and de novo computational protein
Acknowledgements
This work was supported by National Key Research and Development Program of China (2018YFA0901800 and 2018YFA0901400) and National Natural Science Foundation of China (21978015, 21636001, and 21776008).
References (55)
- et al.
A roadmap to engineering antiviral natural products synthesis in microbes
Curr. Opin. Biotechnol.
(2020) - et al.
Engineering the shikimate pathway for biosynthesis of molecules with pharmaceutical activities in E. coli
Curr. Opin. Biotechnol.
(2016) - et al.
Biotechnological production of aromatic compounds of the extended shikimate pathway from renewable biomass
J. Biotechnol.
(2017) - et al.
Microbial synthesis of pyrogallol using genetically engineered Escherichia coli
Metab. Eng.
(2018) - et al.
High-level de novo biosynthesis of arbutin in engineered Escherichia coli
Metab. Eng.
(2017) - et al.
De novo biosynthesis of gastrodin in Escherichia coli
Metab. Eng.
(2016) - et al.
Rewiring carbon metabolism in yeast for high level production of aromatic chemicals
Nat. Commun.
(2019) - et al.
Stepwise modular pathway engineering of Escherichia coli for efficient one-step production of (2S)-pinocembrin
J. Biotechnol.
(2016) - et al.
Engineering 4-coumaroyl-CoA derived polyketide production in Yarrowia lipolytica through a β-oxidation mediated strategy
Metab. Eng.
(2020) - et al.
Engineering the oleaginous yeast Yarrowia lipolytica for high-level resveratrol production
Metab. Eng.
(2020)
Improvement of catechin production in Escherichia coli through combinatorial metabolic engineering
Metab. Eng.
Convergent engineering of syntrophic Escherichia coli coculture for efficient production of glycosides
Metab. Eng.
Balancing the non-linear rosmarinic acid biosynthetic pathway by modular co-culture engineering
Metab. Eng.
Metabolic engineering of Escherichia coli for microbial synthesis of monolignols
Metab. Eng.
Quorum-sensing linked RNA interference for dynamic metabolic pathway control in Saccharomyces cerevisiae
Metab. Eng.
Engineering of L-tyrosine oxidation in Escherichia coli and microbial production of hydroxytyrosol
Metab. Eng.
Synthesis and characterization of hydroquinone glucoside using Leuconostoc mesenteroides dextransucrase
Enzym. Microb. Technol.
Microbial coculture for flavonoid synthesis
Trends Biotechnol.
Heterologous biosynthesis of natural product naringenin by co-culture engineering
Synth. Syst. Biotechnol.
Production of pyranoanthocyanins using Escherichia coli co-cultures
Metab. Eng.
De novo biosynthesis of complex natural product sakuranetin using modular co-culture engineering
Appl. Microbiol. Biotechnol.
Regulating malonyl-CoA metabolism via synthetic antisense RNAs for enhanced biosynthesis of natural products
Metab. Eng.
Production of chemicals using dynamic control of metabolic fluxes
Curr. Opin. Biotechnol.
Coupling feedback genetic circuits with growth phenotype for dynamic population control and intelligent bioproduction
Metab. Eng.
Coupling metabolic addiction with negative autoregulation to improve strain stability and pathway yield
Metab. Eng.
High-throughput screening technology in industrial biotechnology
Trends Biotechnol.
Improving key enzyme activity in phenylpropanoid pathway with a designed biosensor
Metab. Eng.
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The authors contribute equally to this work.