Optimization of Heterologous Glucoraphanin Production <i>In Planta</i>

Barnum, Collin R; Endelman, Benjamin J; Ornelas, Izaiah J.; Pignolet, Roxanna M; Shih, Patrick M.

doi:10.1021/acssynbio.2c00030

Cited by 7 publications

(4 citation statements)

References 37 publications

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“…LC-MS/MS analysis was performed as described by Barnum et al 40 with some modifications. Standard curves were generated using glucoraphanin, glucobrassicin, 4-methoxyglucobrassicin, gluconasturtiin, and sulforaphane for quantification.…”

Section: Methodsmentioning

confidence: 99%

Engineering Brassica Crops to Optimize Delivery of Bioactive Products Postcooking

Barnum,

Cho,

Markel

et al. 2024

ACS Synth. Biol.

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Glucosinolates are plant-specialized metabolites that can be hydrolyzed by glycosyl hydrolases, called myrosinases, creating a variety of hydrolysis products that benefit human health. While cruciferous vegetables are a rich source of glucosinolates, they are often cooked before consumption, limiting the conversion of glucosinolates to hydrolysis products due to the denaturation of myrosinases. Here we screen a panel of glycosyl hydrolases for high thermostability and engineer theBrassica crop, broccoli (Brassica oleracea L.), for the improved conversion of glucosinolates to chemopreventive hydrolysis products. Our transgenic broccoli lines enabled glucosinolate hydrolysis to occur at higher cooking temperatures, 20 °C higher than in wild-type broccoli. The process of cooking fundamentally transforms the bioavailability of many health-relevant bioactive compounds in our diet. Our findings demonstrate the promise of leveraging genetic engineering to tailor crops with novel traits that cannot be achieved through conventional breeding and improve the nutritional properties of the plants we consume.

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Section: Methodsmentioning

confidence: 99%

Engineering Brassica Crops to Optimize Delivery of Bioactive Products Postcooking

Barnum,

Cho,

Markel

et al. 2024

ACS Synth. Biol.

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“…For transient expression, candidate glycosyltransferases were PCR amplified and cloned into the binary vector, PMS057 42 , using Golden Gate assembly 43 , Gibson assembly 44 , or restriction-ligation. XL1-blue Escherichia coli cells were transformed with the assembled plasmids via heat shock 45 .…”

Section: Methodsmentioning

confidence: 99%

Plant-based production of diverse human milk oligosaccharides

Barnum,

Paviani,

Couture

et al. 2023

Preprint

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Human milk oligosaccharides (HMOs) are a diverse class of carbohydrates that aid in the health and development of infants. The vast health benefits of HMOs have made them a commercial target for microbial production; however, producing the ~130 structurally diverse HMOs at scale has proven difficult. Here, we produce a vast diversity of HMOs by leveraging the robust carbohydrate anabolism of plants. This diversity includes high value HMOs, such as lacto-N-fucopentaose I, that have not yet been commercially produced using state-of-the-art microbial fermentative processes. HMOs produced in transgenic plants provided strong bifidogenic properties, indicating their ability to serve as a prebiotic supplement. Technoeconomic analyses demonstrate that producing HMOs in plants provides a path to the large-scale production of specific HMOs at lower prices than microbial production platforms. Our work demonstrates the promise in leveraging plants for the cheap and sustainable production of HMOs.

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“…For transient expression, the native sequences for candidate glycosyltransferases were PCR amplified and cloned into the binary vector, PMS057 (ref. 43), using Golden Gate assembly 44 , Gibson assembly 45 or restriction-ligation (see Supplementary Table 5 for sequences). XL1-blue E. coli cells were transformed with the assembled plasmids via heat shock 46 .…”

Section: Plasmid Construction and Transient Expressionmentioning

confidence: 99%

Engineered plants provide a photosynthetic platform for the production of diverse human milk oligosaccharides

Barnum,

Paviani,

Couture

et al. 2024

Nat Food

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Human milk oligosaccharides (HMOs) are a diverse class of carbohydrates which support the health and development of infants. The vast health benefits of HMOs have made them a commercial target for microbial production; however, producing the approximately 200 structurally diverse HMOs at scale has proved difficult. Here we produce a diversity of HMOs by leveraging the robust carbohydrate anabolism of plants. This diversity includes high-value and complex HMOs, such as lacto-N-fucopentaose I. HMOs produced in transgenic plants provided strong bifidogenic properties, indicating their ability to serve as a prebiotic supplement with potential applications in adult and infant health. Technoeconomic analyses demonstrate that producing HMOs in plants provides a path to the large-scale production of specific HMOs at lower prices than microbial production platforms. Our work demonstrates the promise in leveraging plants for the low-cost and sustainable production of HMOs.

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Optimization of Heterologous Glucoraphanin Production In Planta

Cited by 7 publications

References 37 publications

Engineering Brassica Crops to Optimize Delivery of Bioactive Products Postcooking

Engineering Brassica Crops to Optimize Delivery of Bioactive Products Postcooking

Plant-based production of diverse human milk oligosaccharides

Engineered plants provide a photosynthetic platform for the production of diverse human milk oligosaccharides

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