Molecular and biochemical characterization of a benzenoid/phenylpropanoid meta/para-O-methyltransferase from Rauwolfia serpentina roots

Gomes

Front. Bioeng. Biotechnol.

2020

Curcuminoids are well-known for their therapeutic properties. However, their extraction from natural sources is environmentally unfriendly, expensive and limited by seasonal variability, highlighting the need for alternative production processes. We propose an optimized artificial biosynthetic pathway to produce curcuminoids, including curcumin, in Escherichia coli. This pathway involves six enzymes, tyrosine ammonia lyase (TAL), 4-coumarate 3-hydroxylase (C3H), caffeic acid O-methyltransferase (COMT), 4-coumarate-CoA ligase (4CL), diketide-CoA synthase (DCS), and curcumin synthase (CURS1). Curcuminoids pathway was divided in two modules, the first module included TAL, C3H and COMT and the second one 4CL, DCS and CURS1. Optimizing the first module of the pathway, from tyrosine to ferulic acid, enabled obtaining the highest ferulic acid titer reported so far (1325.1 µM). Afterward, ferulic acid was used as substrate to optimize the second module of the pathway. We achieved the highest concentration of curcumin ever reported (1529.5 µM), corresponding to a 59.4% increase. Subsequently, curcumin and other curcuminoids were produced from tyrosine (using the whole pathway) in mono-culture. The production increased comparing to a previously reported pathway that used a caffeoyl-CoA O-methyltransferase enzyme (to convert caffeoyl-CoA to feruloyl-CoA) instead of COMT (to convert caffeic to ferulic acid). Additionally, the potential of a co-culture approach was evaluated to further improve curcuminoids production by reducing cells metabolic burden. We used one E. coli strain able to convert tyrosine to ferulic acid and another able to convert the hydroxycinnamic acids produced by the first one to curcuminoids. The co-culture strategies tested led to 6.6 times increase of total curcuminoids (125.8 µM) when compared to the mono-culture system. The curcuminoids production achieved in this study corresponds to a 6817% improvement. In addition, by using an inoculation ratio of 2:1, although total curcuminoids production decreased, curcumin production was enhanced and reached 43.2 µM, corresponding to an improvement of 160% comparing to mono-culture system. To our knowledge, these values correspond to the highest titers of curcuminoids obtained to date. These results demonstrate the enormous potential of modular co-culture engineering to produce curcumin, and other curcuminoids, from tyrosine.

Section: Resultsmentioning

confidence: 99%

A Combinatorial Approach to Optimize the Production of Curcuminoids From Tyrosine in Escherichia coli

Gomes

Front. Bioeng. Biotechnol.

2020

“…For cardiovascular diseases, which involve multiple signaling pathways, TCM has its unique advantages. For example, Rauwolfia serpentina root was clinically proven to treat hypertension, fever, liver disease, and other diseases in the past (Wiens and De Luca, 2016;Dong et al, 2018). Besides, current research also proved its good effect on MI/R injury.…”

Section: Acute Myocardial Infarctionmentioning

confidence: 66%

Targeting Calcium Homeostasis in Myocardial Ischemia/Reperfusion Injury: An Overview of Regulatory Mechanisms and Therapeutic Reagents

Wang

et al. 2020

Front. Pharmacol.

Calcium homeostasis plays an essential role in maintaining excitation-contraction coupling (ECC) in cardiomyocytes, including calcium release, recapture, and storage. Disruption of calcium homeostasis may affect heart function, leading to the development of various heart diseases. Myocardial ischemia/reperfusion (MI/R) injury may occur after revascularization, which is a treatment used in coronary heart disease. MI/R injury is a complex pathological process, and the main cause of increased mortality and disability after treatment of coronary heart disease. However, current methods and drugs for treating MI/R injury are very scarce, not ideal, and have limitations. Studies have shown that MI/R injury can cause calcium overload that can further aggravate MI/R injury. Therefore, we reviewed the effects of critical calcium pathway regulators on MI/R injury and drew an intuitive diagram of the calcium homeostasis pathway. We also summarized and analyzed calcium pathway-related or MI/R drugs under research or marketing by searching Therapeutic Target and PubMed Databases. The data analysis showed that six drugs and corresponding targets are used to treat MI/R injury and involved in calcium signaling pathways. We emphasize the relevance of further detailed investigation of MI/R injury and calcium homeostasis and the therapeutic role of calcium homeostasis in MI/R injury, which bridges basic research and clinical applications of MI/R injury.

“…Enzymes involved in lignin precursor formation and those O -methylating the bulky stilbene resveratrol [ 56 ] occupy separate branches. Though several OMTs can methylate more than one substrate [ 57 , 58 ], the main principal reaction is likely maintained within each branch. The alignment underscores a profound impact of the primary sequence of OMTs on their use of aromatic substrates with substrate specificity for O -methylation of hydroxyl groups on aromatic rings favouring the ortho (1,2-) dihydroxy (methoxy) configuration.…”

Section: Discussionmentioning

confidence: 99%

A novel O-methyltransferase Cp4MP-OMT catalyses the final step in the biosynthesis of the volatile 1,4-dimethoxybenzene in pumpkin (Cucurbita pepo) flowers

Hoepflinger,

Barman,

Dötterl

et al. 2024

BMC Plant Biol

Background Floral scents play a crucial role in attracting insect pollinators. Among the compounds attractive to pollinators is 1,4-dimethoxybenzene (1,4-DMB). It is a significant contributor to the scent profile of plants from various genera, including economically important Cucurbita species. Despite its importance, the biosynthetic pathway for the formation of 1,4-DMB was not elucidated so far. Results In this study we showed the catalysis of 1,4-DMB in the presence of 4-methoxyphenol (4-MP) by protein extract from Styrian oil pumpkin (Cucurbita pepo) flowers. Based on this finding, we identified a novel O-methyltransferase gene, Cp4MP-OMT, whose expression is highly upregulated in the volatile-producing tissue of pumpkin flowers when compared to vegetative tissues. OMT activity was verified by purified recombinant Cp4MP-OMT, illustrating its ability to catalyse the methylation of 4-MP to 1,4-DMB in the presence of cofactor SAM (S-(5′-adenosyl)-L-methionine). Conclusions Cp4MP-OMT is a novel O-methyltransferase from C. pepo, responsible for the final step in the biosynthesis of the floral scent compound 1,4-DMB. Considering the significance of 1,4-DMB in attracting insects for pollination and in the further course fruit formation, enhanced understanding of its biosynthetic pathways holds great promise for both ecological insights and advancements in plant breeding initiatives.