Reactive oxygen species (ROS) are not completely “neutralised” oxygen molecules: they still have a “hole” where the electron should be, so they are extremely reactive – to fill this hole. They are often thought about as molecules that oxidise membranes and proteins, and cause ageing. But according to the latest data, ROS are more than just a horror story.
Molecules that oxidise are required in the body to burn glucose, and to form ATP, but it’s impossible to directly turn glucose into ATP, so a lot of processes occur during its breakdown. To understand the value of ROS, we need to understand a bit about these processes. These processes result in the formation of carbon dioxide, as well as special molecules, electron carriers. ATP is synthesised on the mitochondrial membrane in a system called the electron transport chain (ETC).
The ETC is quite complex, but at the same time incredibly logical and beautiful: an aqueduct of several protein complexes is built inside the double membrane of mitochondria. These proteins accept an electron from a carrier molecule and then throw it from one complex to another, like children throwing a ball to each other. Along with the electron transfer, protons are pumped into the double membrane, a strong difference in their concentration on different sides of the membrane – the membrane potential – is formed. At the end of the ETC is an ATP synthase, through which protons are driven to set this machine in motion. But where to put the electrons also traveling along the ETC? This is where oxygen comes in: it accepts them and is reduced to water.
It is the mitochondria, or rather the electron transport chain, that are responsible for the majority of ROS produced. Normally, 90% of the oxygen is reduced to water. But sometimes something goes wrong, and the oxygen is not completely reduced – and the molecule ends up with one (an unstable superoxide radical is formed), two (then peroxide is formed), or three additional electrons (a rarer situation of peroxide reduction to a hydroxyl radical) instead of the four that would be needed to produce water.
Superoxide has a very short lifespan and is not very dangerous. Concentrated peroxide is quite toxic; it is a good oxidiser which means it’s very reactive. Reactivity can cause damage in the cell. It is peroxide that is most often found in the body of mammals due to its stability and long lifespan. Normally, peroxide is constantly present in the cell and, to maintain this level, the coordinated work of both the producing and utilising systems is needed. Peroxide is also able to travel significant distances. Finally, peroxide can take another electron, and then the most toxic compound among the active brethren is formed – the hydroxyl radical. It also lives for a very short time, but is very reactive.
Dangerous or… ?
The free radical theory appeared in the 1960s and has only been developing since then. ROS oxidise (react with) proteins, lipids, and membranes. Errors accumulate in DNA… The consequence of excessive formation of ROS in the cell is indeed oxidative stress. ROS also participate in lipid peroxidation and form hydroperoxides: when they disintegrate, they lead to the emergence of toxic secondary oxidation products.
Peroxide causes oxidation of sulphur-containing groups in the active centres of proteins. It is also associated with the development of oxidative stress and apoptosis (controlled cell death). Oxidative stress in humans is associated with a huge list of various diseases, such as Alzheimer’s disease, Parkinson’s disease, diabetes, atherosclerosis. Of course, oxidative stress is also associated with ageing. Oxidative stress also occurs during brain damage.
At the same time, several independent antioxidant systems coexist in eukaryotic cells. Enzymatic systems include proteins that directly destroy ROS: for example, Mn-containing superoxide dismutase converts two molecules of superoxide into peroxide and water. The peroxide is then decomposed by an enzyme, into water and oxygen. There are also assistant molecules that allow themselves to be oxidised in the name of saving more important structures. The most famous molecule is ascorbic acid. Other known antioxidants are carotenoids — the same pigments that give carrots their hue.
It is easy to think that the appearance of ROS in a cell is just an artifact, a random error. But what if ROS are needed for some reason? This idea is a bit unusual, but it seems to be true, and we need to stop demonising ROS. First of all, it is assumed that ROS can play the role of secondary messengers — they help to carry signals.
Superoxide anions can be used by cells of the immune system to protect against infections and tumours. The fact is that during phagocytosis by macrophages and neutrophils of the immune system, an oxidative explosion occurs: quite a lot of superoxide is formed, triggering oxidative stress in bacteria. Our immune system uses ROS as protection. In addition to its protective function, superoxide is necessary for the synthesis of some important substances, for example, prostaglandins which help with inflammation during injury.
Hydrogen peroxide is the most likely contender for the title of “necessary ROS”. Even the fact that it is formed in the body with enviable regularity hints at its “normality”. There are even specialised structures – microsomes and peroxisomes – in which peroxide forms, too, and oxidative stress occurs.
Peroxide, apparently, takes part in the signaling pathways of both animal and plant cells. For example, the production of pigment by young melanocytes is apparently completely regulated by the release of peroxide by mature cells. In plant cells, peroxide has a role in in the differentiation of xylem. Peroxide even helps tardigrades fall into their suspended animation state of cryptobiosis! Under stress conditions, the tardigrade actively produces ROS, which binds to sulphur-containing cysteine and reversibly changes protein structures.
How can we prove whether a molecule is needed or not? One of the important features of any secondary messenger is the presence of specific targets. And peroxide has such. In particular, tyrosine phosphatases, enzymes that destroy phosphate groups associated with tyrosine residues, change their activity under the influence of peroxide. It oxidises the residues of the cysteine in the active centre, and the proposed cascade has a rather complex regulation and many different participants.
ROS are also involved in much more subtle processes – for example, in the formation of memory. Neurons are connected to each other by axons and dendrites, along which they transmit nerve impulses. Memory, in essence, is certain neural connections, like a road map linking different cities together. And ROS, apparently, quite strongly influence the speed on these roads – how quickly the signal is transmitted.
In addition, peroxide is also involved in gene expression. ROS is able to activate the transcription factor NF-kB, which is necessary for the proper functioning of the immune system; without activation the factor cannot work!
But there is also something that does not quite fit into the theory of the need for ROS. First of all, we still do not understand how exactly their concentration is controlled. There are utilisation systems, but there are also systems that produce ROS randomly. If we assume that ROS are signaling molecules and regulatory cascades exist, then all stages of this process should be catalysed, as occurs in classical regulation. However, it is currently unknown whether there is a controlled source of ROS formation.
ROS are certainly extremely interesting: they cause oxidative stress, indirectly influencing the development of diseases, and, at the same time, are important for normal physiology – or at least so it seems to us for now. Their full role has yet to be discovered.
