For those interested in the science behind these findings: by comparing the cell lysis of untreated and treated bacteria, it is possible to see which proteins have changed. Cell lysis contains "traffic light proteins," proteins involved in various bacterial signaling pathways. By using magnetic beads conjugated to kinase inhibitors, activated proteins can be selected by the available ATP binding sites (green traffic lights). This provides insights into which cellular processes are active or inactivated, providing valuable information about the cell's response to antibiotics. Amanda Holdstad Singleton.
Researchers at the Norwegian University of Science and Technology (NTNU) have developed a promising antibiotic candidate against MRSA. Behind this discovery lies a method that could be important in terms of antimicrobial resistance.
Antimicrobial resistance is a major problem and it's really great to be able to help solve it," said Amanda Holstad Singleton, PhD candidate at NTNU.
Singleton is the lead author of a study that showed how the combination of two new substances was effective in killing methicillin-resistant Staphylococcus aureus (MRSA).
These substances were developed at NTNU and could become a completely new antibiotic that is effective against a wide range of bacteria.
It's one thing to develop new antibiotic candidates, which, when combined, are shown to be well tolerated to human cells, but it's just as important to develop a technology to study how antibiotics work within bacterial cells," Singleton said.
To be able to analyze how these two substances work, the NTNU research team developed a method to analyze how bacterial signaling proteins respond to **. The method provides researchers with a completely new tool for finding new antibiotic candidates.
Up to 10,000 proteins can be found inside bacterial cells. Instead of studying all of these proteins, we "fished out" about 2,000 or so proteins, which are signaling proteins. These proteins control most of what happens in cells," Singleton said.
This method allows researchers to see if each of these proteins is activated or inactivated after adding the substance they want to test.
These proteins can be likened to a traffic light, which can change from red to green and back again. By turning them red, you stop the vital signaling pathways inside the cell," Singleton says.
If a substance is found to affect signaling proteins by switching the traffic lights of key processes within the cell to red, it is considered a candidate for a new antibiotic. If a substance is found to produce red light in several different processes in the cell, then it is a better candidate.
This is exactly what NTNU researchers have done after combining two different substances that could become new antibiotics.
In a recent study published in the journal Frontiers in Microbiology, we found that the combination of two new substances developed by NTNU was more effective at killing MRSAs than when used alone," Singleton said.
About four years ago, researchers from the Department of Clinical and Molecular Medicine at NTNU published the bactericidal properties of a specific type of peptide. These peptides, combined with compounds developed in the Department of Chemistry of NTNU, may now be a completely new antibiotic.
Peptides are chains of amino acids, and amino acids are the building blocks of proteins. What is special about these particular peptides is that they bind to proteins in bacteria, which is absolutely necessary for bacteria to be able to replicate their DNA," says Professor Marit Otterlei.
Peptides prevent DNA from replicating and hence the bacteria die.
No other antibiotic attacks this protein. This means that it is a new target, so no bacteria are resistant to these peptides. Since this target protein is present in all bacteria, these peptides also act against multidrug-resistant bacteria," Otterlei said.
While Otterlei and her colleagues continue to work on these peptides, researchers Eirik Sundby and BrD Helge Hoff from the Department of Materials Science and Engineering and the Department of Chemistry at NTNU are working to find substances that effectively block the formation of DNA building blocks. They also developed a compound known as a kinase inhibitor that can be used to fish out signaling proteins from bacterial samples.
When the method was ready, we tested it on bacteria treated with peptides, one of these new molecules that is thought to influence the production of DNA building blocks. We found that the new molecules have a different mechanism of action than we thought, but they do have a very good binding effect with our peptides, the so-called synergistic effect," Otterlei said.
It turns out that the new molecule developed by Sundby and Hoff inhibits energy metabolism within bacterial cells. Binding to the peptides of Otterlei, they also lead to the activation of proteins associated with multiple stress responses in bacterial cells. This does not happen when these substances are administered alone. This extra activation results in the bacteria dying more efficiently.
According to the researchers, this is the first time that the effectiveness of antibiotics has been studied in this way.
This gives us a completely new way to evaluate new antibiotic candidates," Otterlei said.
It also provides researchers with a new way to prevent resistance to new antibiotics.
We must remember that the development of drug resistance is a natural part of evolution. It's inevitable. However, for bacteria, developing resistance is costly. It has to make some sacrifices," Otterlei said.
Singleton explained that bacteria can become resistant to antibiotics in two ways: either through contact with bacteria that are already resistant to antibiotics and exchanging DNA between them, or by mutations in the bacteria's genes that protect it from antibiotics.
This type of mutation comes at a cost, and it affects the adaptability of bacteria. One trait is sacrificed for the sake of another trait that provides antibiotic protection," Singleton said.
If the benefits of being protected from antibiotics outweigh the disadvantages, bacteria will multiply and we will get many new antibiotic-resistant bacteria.
However, this job becomes more difficult if the bacteria have to develop resistance to two substances at the same time, both of which work in completely different places within the bacterial cell.
If you attack two different processes, it will be too much of a burden to develop resistance to both, and the bacteria will become less viable.
If you also create a way for antibiotics to attack bacteria to develop resistance, it becomes even more difficult.
In our case, the protein attacked by our new antibiotic candidate plays such a critical role in replicating the bacteria's DNA that if a mutation occurs, the loss of adaptability becomes so great that the bacteria die," Singleton said.
More information: Amanda Holstad Singleton et al, Activation of multiple stress responses in Staphylococcus aureus significantly reduces minimum inhibitory concentrations when combining two novel antibiotic candidates, Frontiers in Microbiology (2023). doi： 10.3389/fmicb.2023.1260120