Chuck Bednar for redOrbit.com – @BednarChuck
People who are adept at homebrewing beer may soon be able to brew their own antibiotics, anti-cancer medication, or other drugs thanks to a due technique discovered by bioengineers from the University of California, Berkeley and an international team of colleagues.
In research published Monday in the journal Nature Chemical Biology, John Dueber, an assistant professor of bioengineering at UC Berkeley, and his fellow researchers explained how they built on the homebrewing technique. The technique consists of using yeast to convert sugar into alcohol and creating a microbial factory capable of producing drugs.
In a statement, they explained that they focused on replicating a complex, 15-step process in poppy plants in microbes – a process that allows production of therapeutic substances. While previous efforts have been able to recreate different portions of the poppy’s drug pathway using yeast or E. coli, Dueber’s team for the first time was able to replicate the final steps, enabling the task to be completed from start to finish in a single organism.
They replicated the early steps in the process using an engineered strain of yeast, and then they were able to synthesize a compound found in poppy called reticuline from a glucose derivative known as tyrosine. The authors said that their research is the first to describe each step of the process, from feeding yeast glucose through the synthesis of the target drug, and that the challenge now is to link each step together and to work on scaling-up the process.
Bridging the gap in the poppy pathway
The researchers explained that the poppy pathway is an attractive target for such studies because it contains a highly active class of bioactive compounds known as Benzylisoquinoline alkaloids (BIAs). This family of natural products contains some 2,500 molecules that can be used for such bioactivities as painkillers, antibiotics, and anti-cancer therapeutics, Dueber told redOrbit.
Yeast cells producing the yellow beet pigment betaxanthin, which UC Berkeley researchers used to quickly identify key enzymes in the production of benzylisoquinoline alkaloids (BIAs). Credit: Berkeley
The part of the pathway in which the glucose is converted to reticuline, “the last intermediate common to all types of these BIAs” and a “major branch-point hub,” had previously only been accomplished in E. coli, he explained via email. The downstream enzymes of most of these branches, however, had only previously been successful using yeast “because they contain enzymes that are notoriously problematic to functionally express in E. coli.”
“Ultimately, you want the entire pathway for producing your desired molecule of interest to all be expressed in the same cell because this can be robustly grown in large-scale fermenters,” the UC Berkeley bioengineer told redOrbit. His team isolated then improved an enzyme that completes the first missing step in the pathway in yeast, and then extends it to produce reticuline.
“We accomplished this by expressing a flower (Mirabilis jalapa) enzyme that converts the product of our missing enzymatic step (L-DOPA) into a molecule that is highly fluorescent and colored,” he said. “This allowed us to both isolate a beet enzyme capable of doing this reaction (the enzyme is involved in synthesizing the violet pigment that gives beets their color) and then further improving the enzyme by mutating its sequence and picking the cells that were the most fluorescent/colored. We then replaced this flower enzyme for synthesizing the fluorescent biosensor with the downstream BIA producing enzymes.”
Previous research demonstrated that reticuline fed to yeast cells could be converted to codeine, and another study has demonstrated the conversion of of thebaine to morphine in yeast. The steps featured in those studies overlap, establishing the steps to go from reticuline to morphine, Dueber said. One last step is required to convert connect all of the dots – epimerization from one type of reticuline to the other – and he is confident that another lab will soon describe this process.
Several possible benefits, but concerns as well
Dueber told redOrbit via email that there are “several potential benefits” to their research. For instance, it “enables the production of this large family of natural products, hopefully in high concentrations once the before mentioned inefficient enzymatic steps are improved. Again, many of these have potential for a variety of desirable bioactivities, but we are currently limited in studying many since the natural plants make them in vanishingly low amounts.”
“Yeast are much easier to genetically reprogram than plants and also replicate much faster,” doubling their population in about two hours, he added. “This should allow us to introduce non-native enzymes to make so-called unnatural natural molecules, further increasing the diversity of this already large BIA family. We may be able to make therapeutics that have even more desirable bioactivity.”
This could allow them to produce a steady supply of morphine that is not subject to weather conditions, pests, or other destructive elements. The supply of the substance could be adjusted to match the medical field’s demand, Dueber explained, and the fermentation process could be done in a secured location. He and his colleagues believe that sugar-fed yeast could reliably produce these substances in only a few years’ time, and they are calling for regulators and law enforcement officials to take note of their research.
“There are considerable challenges to be overcome, but I think a morphine-producing strain could be achieved in a two-to-three-year timeframe if it was made the central focus of a talented metabolic engineering lab given the rapid progress in the last couple of years,” he told redOrbit via email. “Due to the concern of the illicit potential misuse of such a strain, the authors of our paper think it is extremely important to engage the public and government now to consider the most sensible steps to take to encourage the research that holds great potential for benefiting society while taking measures to make the illicit uses as difficult as possible.”
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