Strychnine, the highly toxic alkaloid found in vomit, not only inspires crime writers. Since the discovery of the toxin, chemists have also been fascinated by its complex molecular structure. Although strychnine can be chemically synthesized for decades, it has been unclear how nature produces this extraordinary toxin. Researchers have now traced the synthesis pathway in the crushed nut in detail. They transferred the genes involved into a tobacco plant, which then also produced strychnine.
In Agatha Christie’s debut work The Missing Link in the Chain, the victim dies, convulsing, of strychnine poisoning. The poisonous alkaloid of the vomit (Strychnos nux vomica), which has long been used as a rat poison, is also used in numerous other crime novels and films. The poison triggers deadly muscle cramps even in small doses. Strychnine was chemically isolated for the first time in 1818, and in 1946 the chemist Robert Robinson succeeded in elucidating the complex structure of the molecule. Eight years later, Robert Burns Woodward developed a process to chemically synthesize strychnine. The way in which strychnine is produced in nature in the nutmeg has long been a mystery to science.
Search for clues in the genome
A team led by Benke Hong from the Max Planck Institute for Chemical Ecology in Jena has now unraveled this puzzle. “Our key question was how we can find the genes responsible for the biosynthesis of strychnine in the vomit,” says Hong. “In a first step, we compared the expression of the genes of two species of the same genus (Strychnos), of which only the vomit tree produces strychnine.”
Based on this, they identified genes that may be involved in the production of strychnine. In doing so, they orientated themselves on assumptions about chemical conversion processes that proceed via several intermediate products. One of these intermediate products, geissoschizin, is also produced in the medicinal plant Madagascar periwinkle, among other things. The steps leading to this intermediate product have already been biochemically elucidated – and Hong and his colleagues did indeed find the corresponding genes in the vomit genome.
Chemical detective work
For the next steps, the researchers were guided by chemical considerations: “Based on the chemical structures and mechanisms, a proposal for the chemical conversion emerged for each step in the metabolic pathway,” explains Hong’s colleague Sarah O’Connor. For each chemical reaction from intermediate to intermediate, they thought about what kind of enzyme might be needed and then looked for a corresponding gene.
In order to check whether the genes found actually have the assumed task, the researchers transferred them to tobacco plants, which then also produced the respective enzymes. If the researchers then added the corresponding starting materials, it was shown whether these were converted by the enzymes into the next products of the synthesis path. In this way, the team identified all the genes leading up to the production of the molecule prestrychnine, the final precursor of strychnine.
discovery by chance
They could only find no enzyme for the last step in the synthesis, the conversion of prestrychnine into strychnine. “We initially thought that this process must be catalyzed by one or more enzymes. In fact, we examined many enzymes, but none of them were reactive,” reports Hong. But coincidence came to the researchers’ aid: “Surprisingly, one day I found that a sample of prestrychnine that had been stored at room temperature on the laboratory bench had slowly converted to strychnine over time,” says Hong. Apparently, the last step does not require any enzymes, but takes place spontaneously.
In addition to the biosynthetic pathway of strychnine, the researchers also found out how related molecules are produced – brucine in the vomit and diabolin in another species in the genus Strychnos. The results now make it possible not only to synthesize the corresponding molecules chemically, but also to produce them using a process called metabolic engineering. In the process, individual enzymes are modified in natural metabolic pathways in such a way that the desired product is formed. Among other things, this makes it easier to obtain active ingredients for medical applications.
Source: Benke Hong (Max Planck Institute for Chemical Ecology, Jena) et al., Nature, doi: 10.1038/s41586-022-04950-4