Chemists modify E. coli genetic code to produce 21st amino acid, giving new capabilities

State University chemist Han Xiao and his team have successfully expanded the genetic code of Escherichia coli bacteria to produce a synthetic building block, a “noncanonical amino acid.” The result is a living indicator of oxidative stress.

The work, they say, is a step towards technologies that can enable the generation of new proteins and organisms with a variety of useful functions.

Their study appears in the journal Cell Press Chem.

Amino acids are the building blocks of DNA. In general, organisms only need 20 to program the entire set of proteins needed for life. But Xiao, with the support of a $ 1.8 million grant from the National Institutes of Health, set out to see how a 21st amino acid would enable the design of ‘unnatural organisms’ that serve specific purposes.

The new study does just that by engineering bacteria to produce the extra amino acid, called 5-hydroxyl-tryptophan (5HTP), which appears naturally in humans as a precursor to the neurotransmitter serotonin, but not in E. coli. The new production of 5HTP requires the bacteria to produce a protein that fluorinates when the organism is under metabolic stress.

“The process requires a lot of interdisciplinary techniques,” Xiao said. “In this study, we combine synthetic chemistry, synthetic biology and metabolic engineering to create a strain that synthesizes and encodes a 21st noncanonical amino acid, and then we use it to produce the desired protein.”

Xiao said that programming the autonomous unnatural bacteria was a three-step process: First, the researchers, led by graduate student Yuda Chen, created bioorthogonal translation machines for the amino acid, 5HTP. Second, they found and targeted a low codon – a sequence in DNA as RNA that does not produce protein – and genetically engineered to encode 5HTP. Third, by degrading enzyme clusters of other species in E. coli, they gave the bacterium the ability to produce 5HTP.

“These 5HTP-containing proteins, isolated from the programmed bacteria, can be further labeled with drugs than other molecules,” Xiao said. “Here we show that the strain itself can serve as a life indicator for reactive oxygen species, and the detection limit is really low.”

Although researchers have so far reported on the creation of more than 200 non-canonical amino acids, most of them cannot be synthesized by its host organisms. “This has been a continuous field for decades, but rather people focused on the chemical part,” Xiao said. “Our vision is to manipulate whole cells with the 21st amino acid that allows us to investigate biological or medical problems in living organisms, instead of just dealing with cells in the lab.

“Moving from this technology to the host type eliminates the need to inject artificial building blocks into an organism because they can synthesize and use it on their own,” he said. “That allows us to study non-canonical amino acids at a higher level, whole organism.”

Finally, the researchers hope that custom building blocks will allow cells, such as those in tumors, to make their own therapeutic drugs. “That’s an important future direction for my lab,” Xiao said. “We want cells to detect disease, make better medicines and release them in real time. We do not think this is too far away.”

Co-authors of the paper include Rice postdoctoral fellows Juan Tang, Lushun Wang and Zeru Tian, ​​undergraduate student Adam Cardenas and visiting scientist Xinlei Fang, and Abhishek Chatterjee, an assistant professor of chemistry at Boston College. Xiao is the Norman Hackerman-Welch Young Investigator and an Assistant Professor of Chemistry.