Images of proteins involved in the formation of a strong antibiotic reveal the first unusual steps of the synthesis of the antibiotic. The better understanding of the chemistry behind this process, outlined in a new study led by Penn State chemists, could allow researchers to adapt this and similar fertilizers for use in medicine human.
The antibiotic thiostrepton is highly potent against Gram-post pathogens and may even target some breast cancer cells in culture. Although it has been used topically in medicine, so far it has been ineffective in humans because it is badly caught. We studied the first steps in thiostrepton biosynthesis in the hope that we can eventually remove some processes and make analogs of the molecule that may have better pharmacological properties. Importantly, this reaction is found in the biosynthesis of several other antibiotics, so the work has the potential to be extensive. “
Squire Booker, Biochemist, Penn State and Researcher with the Howard Hughes Medical Institute
The first step in thiostrepton synthesis involves a process called methylation. A molecular tag called a methyl group, which is important in many biological processes, is attached to a molecule of tryptophan, the substrate of the reaction. One of the main systems for methylating fertilizers that are not particularly reactive, such as tryptophan, involves a class of enzymes called radical SAM proteins.
US radical proteins typically use an iron-sulfa broth to purify a molecule called S-adenosyl-L-methionine (SAM), which produces a “free radical” or an unrepaired electron. help move the reaction forward, “said Hayley Knox, a graduate student in chemistry at Penn State and first author of the paper.” It’s the only exception we know so far. this is the protein involved in thiostrepton biosynthesis, called TsrM. We wanted to understand why TsrM does not perform radical chemistry, so we used an imaging technique called X-ray crystals to study a structure at several stages during its reaction. “
In all SAM radical scavenging proteins identified so far, SAM binds directly to the iron-sulfa group, which helps break down the molecule to produce the free radical . However, the researchers found that the site where SAM would normally connect is blocked in TsrM.
“This is completely different from any other radical US protein,” Booker said. “Instead, the proportion of SAM that binds to the related groups of the tryptophan substrate and plays a key role in the reaction, in what is known as substrate-assisted catalysis. “
The researchers present their findings in an article appearing Jan. 18 in the journal The chemistry of nature.
In dissolving the structure, the researchers were able to detect the chemical steps in the first part of thiostrepton biosynthesis, when tryptophan is methylated. In a short time, the methyl group from SAM migrates to a part of TsrM called cobalamin. Then, with the help of an additional SAM molecule, the methyl group moves to tryptophan, replenishes free cobalamin and removes the methylated substrate, which is essential for the next steps in synthesizing the antibiotic.
“Cobalamin is the strongest nucleophile in nature, which means it is highly reactive,” Knox said. “But the tryptophan substrate is weakly nucleophilic, so a big question is how cobalamin could be excreted. We found that arginine residue sits under the cobalamin and disinfects the methyl-cobalamin, allowing tryptophan to dissolve cobalamin and become methylated. “
Next the researchers plan to study other cobalt-dependent SAM radical proteins to see if they work in similar ways. Eventually, they hope to find or create analogues of thyostreptone that can be used in human medicine.
“TsrM is clearly unmatched in terms of known SAM radical proteins that are dependent on cobalamin and radical SAM proteins in general,” Booker said. “But there are hundreds of thousands of specific sequences of radical US enzymes, and we still don’t know what most of them do. As we continue to study these proteins, we may there for many more surprises. “
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Magazine Reference:
Knox, HL, et al. (2021) Structural basis for non-radical catalysis with TsrM, SAM radical methylase. Natural Chemical Biology. doi.org/10.1038/s41589-020-00717-y.