Unravelling the role of redox modifications Reactive oxygen species (ROS) can cause oxidative stress and protein damage in organisms, but they are also involved in signalling processes and regulate cellular processes. Professor Haike Antelmann tells us about her research into specific protein modifications caused by ROS, molecular switches that play a role as oxidative stress defense mechanisms in bacteria, and are important for the development of effective treatment against specific pathogens The exposure of bacteria to ROS results in oxidative stress responses and cellular damage, but low doses of ROS are also implicated in signalling processes and regulate specific thiol-switches. Based at the Freie Universität in Berlin, Professor Haike Antelmann is the Principal Investigator of the Mycothiolome project, an ERC-backed initiative investigating the topic. “My project is investigating the role of these thiol-switches, protein modifications that play a role under oxidative stress conditions,” she outlines. When bacteria are exposed to ROS during infection, or just in the course of everyday life due to aerobic growth, specific proteins inside the bacteria are modified by ROS. “ROS can damage proteins, and hence bacteria need specific protection and defense mechanisms,” explains Professor Antelmann. “Low molecular weight (LMW) thiols help to protect against protein damage and to keep the redox balance.”
Low molecular weight Thiols in bacteria This forms a core part of Professor Antelmann’s research agenda. Actinomycetes, specific members of Gram-positive bacteria, are an area of particular interest in the project. “Mycothiol is the major LMW thiol in Actinomycetes which can modify proteins under oxidative stress,” says Professor Antelmann. In the case of eukaryotes and most Gram-negative bacteria, the major LMW thiol is glutathione (GSH), an antioxidant that protects cellular components from damage by ROS. There is a large body of research about the role of protein modifications by GSH in humans, which are implicated in many physiological and pathophysiological processes, and hence can be thought of as molecular switches controlling human health or disease. However, Grampositive bacteria, such as Actinomycetes and Firmicutes do not produce GSH, but instead utilize alternative LMW thiols, such as mycothiol (MSH) and bacillithiol (BSH). “We have found that proteins are protected under oxidative stress by MSH and BSH. These protein modifications can lead to changes in the activity of the proteins, meaning that the proteins become inactive or active, which has a regulatory effect,” continues Professor Antelmann.
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We identified 58 proteins with S-mycothiolations in M. smegmatis under HOCl stress that are shown by color codes based on their % oxidation and classified into different functional categories in this Voronoi treemap. The cell size denotes the protein abundance in the total proteome. [Figure 2 published in Hillion et al., Scientific Reports 7: 1195. (2017)].
The project’s primary focus is fundamental research into the function and structure of specific proteins and how they are modified and what is the physiological consequence for the bacteria. Professor Antelmann and her colleagues in the project are using sophisticated techniques, including mass spectrometry and novel thiol-redox proteomics approaches, to analyse the S-mycothiolome under oxidative
to protect proteins from lethal damage in these conditions.” A number of interesting results have been gained, while researchers are also investigating the process by which proteins are switched-on or off between different conformational and functional states. Thiol switches play a central role in this regard. “We aim to find out what the most important thiol switches are with respect
We are investigating which modifications occur under oxidative stress, and the effect on the physiology of the cells. We have found that proteins are protected and redoxcontrolled under oxidative stress by mycothiol, which could be important under infection-related conditions stress conditions. “In this project we have applied new redox proteomics methods and visualization tools to unravel the different kinds of protein modifications in a quantitative manner. More than a thousand proteins are modified in different ways under oxidative stress conditions,” she says. There are several different forms of oxidative stress; one major area of research is hypochlorous acid stress. “Hypochlorous acid (HOCl) is a very potent oxidant,” explains Professor Antelmann. “Mycothiolations, protein modifications by the LMW thiol MSH, can help
to these protein modifications. With these thiol switches, you can almost switch a protein on or off,” explains Professor Antelmann. Many thiol switches have been found, and researchers now aim to develop a deeper understanding of the underlying mechanisms involved and their wider effects on cellular physiology. “Do these modifications play an important role in protecting the proteins against damage or do they change protein activity? We want to understand the physiology behind these protein modifications,” outlines Professor Antelmann.
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