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Image: In addition to conducting genetic and biochemical studies, the scientists used a green fluorescent protein (GFP) "tag" and microscopy equipment from the Brookhaven Lab's Center for Functional Nanomaterials to visualize the proteins
they studied in leaf cells.
Left: Localization
of GFP-tagged electron donor proteins along the endoplasmic reticulum (the internal network of membranes) in leaf cells.
Right: GFP-labeled P450 enzyme complex interacting
with electron donors.
In this case, the scientists attached half of the GFP tags to these proteins; Fluorescence occurs
only when the two parts of the protein interact.
(In both images, the red signal comes from chlorophyll
.
) )
Source: Brookhaven National Laboratory
Plant biochemists at the U.
S.
Department of Energy's Brookhaven National Laboratory have discovered a new biochemical "mechanism" by which plants convert organic carbon produced by photosynthesis into a series of ring-shaped aromatic molecules
.
The study, which has just been published in the journal Science Advances, proposes new strategies
for controlling plant biochemistry in agricultural and industrial applications.
"Our study reveals the long-overlooked complexity and versatility of a group of key enzymes known as cytochrome P450 monooxygenase," said
Chang-Jun Liu of the Department of Biology at Brookhaven Laboratory.
"These enzymes act like a synthesis machine that produces a wide variety of aromatic compounds in plants, including compounds that build the plant's waterproof backbone and vasculature, as well as compounds
that resist insect invasion and ultraviolet radiation.
"
Revealing the complexity of how these enzymes are regulated gives scientists a new set of genetic tools that can be used to precisely control which compounds are produced
in the body.
This work can help promote long-term carbon storage and carbon-neutral use of plant biomass for energy applications, improve the nutritional properties of plants, or enhance their resistance to disease and harsh environmental conditions
.
Scientists have long known that the P450 enzyme cannot determine the structure and biological characteristics
of aromatic compounds alone.
"In order for the P450 machines to operate, they need partner molecules to transfer electrons
.
These electrons act like a power source that fuels the machine
.
”
Traditionally, scientists thought that P450s interacted primarily with a common electron donor called cytochrome P450 reductase to produce various aromatic compounds
.
But new research shows that different P450s selectively work with different electron donors (and electron transport chains) to drive their activity
.
In addition, the researchers found that the same P450 enzyme can produce different kinds of aromatic hydrocarbons
using different electron donors and electron transport chains in different parts of the plant (stems, leaves and seeds).
The scientists made these discoveries by analyzing aromatic compounds that accumulate in different parts of plants where genes from different electron donors were selectively deleted
.
"By knocking out these genes, we were able to determine the contribution of different electron donors, determine which electron donors drive the production of different aromatic hydrocarbons in different parts of the plant, and then, in yeast cells, we reassembled different electron transport chains in combination with plant P450 enzymes to mimic reactions
in plants.
" These studies helped us validate the contribution
of individual electron donors and transport chains in supporting P450 activity.
”
"Plants have evolved many homologous genes as electron donors, so we need to create plants
with single-gene deletions and gene combination deletions.
" We then studied the changes
in the product distribution of aromatic hydrocarbons during plant growth.
”
Xianhai Zhao said: "We also conducted a comprehensive comparative analysis of electron donor gene expression and electron source molecular abundance in different parts of the plant, and measured the electron transfer rate
of different transport chains.
"
These experiments helped scientists identify potential reasons why
certain P450 enzymes work with different electron transport chains in different parts of the plant.
The knowledge gained provides scientists with a new set of genetic tools that they can manipulate to control aromatic production
.
"We can manipulate a specific electron donor, instead of p450, to inhibit a different set of aromatic hydrocarbons and achieve the desired result.
"
For example, reducing an aromatic compound called lignin in the stem could make it easier for plants to break down and convert them into biofuels
.
Reducing the amount of certain aromatic hydrocarbons in seeds can improve their nutritional value
.
Chang-Jun Liu said: "The detailed knowledge provided in this study allowed us to make selective changes to one part of the plant without affecting another part, such as accumulating aromatic compounds
in the leaves that provide UV sun protection.
"
The Brookhaven team plans to test these genetic manipulation strategies to optimize bioenergy crops
.
They will also conduct further research using cryo-electron microscopy at the Brookhaven Biomolecular Structure Laboratory to understand the atomic-level details
that drive the selective partnership between the P450 enzyme and a specific electron donor.
"Exploring the molecular basis of selective P450-electron donor binding will help us further our understanding of how the P450 system works
.
" "This, in turn, will make it possible for us to create more efficient enzyme systems to produce the desired biological products and enhance carbon conversion and storage
through photosynthesis.
"
Tissue-preferential recruitment of electron transfer chains for cytochrome P450-catalyzed phenolic biosynthesis
