A team of chemists from the University of Tokyo has developed 10 new artificial forms of an old antibiotic called gramicidin A, which appear to be safer, stronger drugs for use in humans.

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The researchers published their findings in the journal Nature Communications.

As bacterial resistance to antibiotics steadily grows, researchers are focusing on finding improved antibacterial compounds. Scientists are constantly working on strategies to enhance and transform natural products to improve human health.

Gramicidin A is one of the world’s oldest antibiotics. Following its discovery in soil bacteria in 1939, gramicidin A became the very first antibiotic to have commercial use. Its initial success paved the way for scientists to develop penicillin and ushered in the antibiotic era.

Currently, doctors prescribe gramicidin A as an ingredient in topical creams or drops to treat infected surface wounds and eye, nose, and throat infections.

It is only suitable for external use because a significant intake may cause the breakdown of red blood cells and can be toxic to organs, such as the kidney and liver.

Gramicidin A kills bacteria by forcing itself through the cell membrane, a layer that physically separates the components of the cell from the outside environment. By doing this, it causes the contents of the cell to leak out and the surrounding contents to leak in through nano-sized tunnels called ion channels.

In this way, gramicidin A destroys bacterial cells. However, it has the same effect on human cells.

The way in which gramicidin A produces these ion channel-like pores has attracted significant interest from the scientific community because ion channels are abundant in nature.

Human ion channels play a wide range of vital roles in the body, from maintaining blood pressure to moderating brain function. The disruption of ion channels negatively influences the ability of cells to control their internal environment.

Scientists have developed approximately 350 artificial versions of gramicidin A over the past 80 years, all of which have properties comparable to the original and are, therefore, unsuitable for use in humans.

In the present study, chemists at the University of Tokyo Graduate School of Pharmaceutical Sciences and Teikyo University Institute of Medical Mycology, both in Japan, designed, constructed, and analyzed almost 4,100 variations of gramicidin A. They hoped to create a better, safer drug.

The scientists report that gramicidin A is a spiral of 15 amino acids. Amino acids are the building blocks of proteins.

In their experiments, the team strategically selected six of these amino acids that can undergo alteration without affecting critical aspects of gramicidin A’s structure.

It was possible to exchange each of those six amino acids with four alternative amino acids. This led to a total of 4,096 variations.

The team employed a “one-bead one-compound” synthesis technique, in which small glass beads serve as the foundation to attach the first amino acid. The researchers assembled the peptide by adding further amino acids one at a time.

“Usually, natural product synthesis is a very difficult, complicated task. There are many steps to make these large molecules, and, at the end, synthetic yields are very low, so synthetic approaches like the one-bead one-compound synthesis we used are still uncommon with natural products,” says Hiroaki Itoh, an assistant professor and co-author of the study.

After completing their synthesis, the scientists put each of the beads into a separate container and evaluated the function of their new variations of gramicidin A.

“Actually, this was a fully manual operation. It was a struggle for the student in charge of the project, but she is a very hard worker and made a great accomplishment with this research. Considering the normal timeline of natural product chemistry, this was quick,” says Itoh.

The student, Yuri Takada, lead author of the research paper, has gone on to complete her doctoral studies and is currently a postdoctoral researcher at the University of Cambridge in the United Kingdom.

Next, the scientists began testing their new variations of gramicidin A for activity against strains of Streptococcus bacteria that are responsible for many bacterial infections. Streptococcal infections cause many diseases, including strep throat and pneumonia.

The scientists then tested the best performing new versions of gramicidin A for safety. They assessed how these interacted with rabbit blood cells and mouse leukemia cells.

Their preliminary laboratory tests identified 10 gramicidin A variations as potential future antibacterial drugs.

The findings also allowed the scientists to determine how specific structural changes to the amino acids affect the overall function of the molecule. They discovered that although all 10 variations share a similar ion channel function, they have different effects on the cells.

The team also evaluated the ion channel-forming ability of these high-performing new versions of gramicidin A. While these versions were less harmful to mammalian cells, their ion channel-forming ability remained strong.

“It has long been believed to be very difficult to realize species-selective ion channel-forming activity, but our study showed gramicidin A can have very bacteria-selective activity,” says Itoh.

The foundational structure-function evidence that the team demonstrated is crucial for understanding why and how pharmaceuticals work.

The authors conclude in their paper that the approach that they developed for this study “will be widely applicable as a promising strategy for identifying key structural features and optimizing the pharmacologically favorable activity of natural products.”