For background information on this post, start by reading the introduction to oixdation. Terms you should understand before reading this article are superoxide and ROS. Additionally, you should understand the basics of the human antioxidant system.
Hydrogen Peroxide is the Signal
Superoxide is a free radical which is produced during metabolism at the bottleneck in the Electron Transport Chain. It is rapidly converted to hydrogen peroxide by an enzyme called superoxide dismutase (SOD). SOD is a fascinating enzyme – it is the fastest enzyme ever discovered. As fast as superoxide can diffuse to SOD, SOD can turn it into hydrogen peroxide. SOD is an ancient enzyme that is present in all branches of life, demonstrating that it evolved before the differentiation of bacteria and archaea. SOD is of truly primal importance.
Hydrogen peroxide is an oxidant but it is not a free radical. It is also a neutral molecule, unlike superoxide which has a negative charge. This gives hydrogen peroxide several useful characteristics. As a small molecule with a neutral charge it can diffuse through cell membranes, meaning that mitochondrially derived hydrogen peroxide can move throughout the cell. It is also less reactive than superoxide. Cells can actually have significant buildup of hydrogen peroxide without leading to the out of control reaction cascades associated with free radical induced oxidative stress, such as lipid peroxidation.
As I have repeated throughout this blog, “ROS is the signal”. Most often hydrogen peroxide is the specific Reactive Oxygen Species delivering the signal. How is the message delivered?
A Very Specific Message
Proteins do most of the work in cells. Enzymes are proteins that act as catalysts, performing functions such as cleaving a bond in a sugar molecule to help oxidize it. Proteins are made of little subunits called amino acids. DNA carries genes, which are just blueprints for how to build proteins. The gene tells the cell the exact sequence of amino acids to put in the protein and the sequence of amino acids determines the function of the protein.
The body has all kinds of ways of regulating proteins. First and foremost, it can decide whether or not to produce the protein that a gene codes. Typically, proteins won’t be produced unless another protein called a “transcription factor” binds to DNA near it in its “promoter” . When the transcription factor binds the promoter, the gene is turned on and the protein is made!
Of the 20ish amino acids that make up proteins, the one most relevant to redox biology is cysteine. According to this paper , “Of the 20 common amino acids, perhaps the most intriguing and functionally diverse is cysteine … is subject to alkylation by electrophiles and oxidation by reactive oxygen and nitrogen species, leading to posttranslationally modified forms that can exhibit significantly
altered functions.” The term “posttranslationally” means “after the protein is made. Just because a protein exists, doesn’t mean that it can do anything yet.
The “thiol group” on the cysteine lends itself to oxidative modification and acts like a lightswitch on a protein. Depending on its oxidative state, the protein’s function might be turned on or off. The oxidative state, and thus whether the protein is active is determined by the amount of hydrogen peroxide in the immediate vicinity of the protein. We can see that these thiol groups are incredibly important because cysteine is one of the most strongly conserved amino acids. This means that as evolution moves on over time, you don’t mess with the part of the protein that has the cysteine! (Actually, when cysteine is in other parts of the protein it is very poorly conserved, as if evolution is trying to get rid of it.)
Let’s put it all together. When superoxide is produced in a cell it is rapidly converted to hydrogen peroxide. This is great, because hydrogen peroxide is not a strong enough oxidant to damage proteins, DNA or fats. Many of the functional proteins in the cells contain crucial cysteine amino acids. The thiol groups on those cysteine subunits can be oxidized by hydrogen peroxide. That oxidation is the switch that turns them on or off.
Got it? Good. Ready for a real world example? Are you sure?
Biology Is Weird
Let’s think about insulin signalling. When you eat a meal, blood sugar rises (depending on what you’ve eaten) and insulin is produced in the pancreas and released into the bloodstream. The insulin binds to an insulin receptor on a cell surface, which is a signal to the cell that glucose is available and the cell should start taking it in and oxidizing it. But at first the insulin receptor is turned off. How does it get turned on?
The insulin receptor turns itself ON through a process known as autophosphorylation – it adds a phosphate group to itself. When insulin levels are low, the receptor is turned off by another enzyme called a phosphatase – whose job it is cut off the phosphate group that the insulin receptor adds to itself to turn itself on. Still following? So the insulin receptor is turned ON when it’s phosphorylated and OFF when it’s not. When insulin binds, it stimulates yet another protein – NADPH oxidase – which releases a short burst of superoxide, which is rapidly converted to hydrogen peroxide.
The phosphatase enzyme, tyrosine phosphatase 1B, the one that REMOVES the phosphate group, is highly prone to being oxidized. Specifically this is done by hydrogen peroxide oxidizing the cysteine located in the active site. If you’re really interested you can read about it here. The burst of peroxide actually turns it off. When that happens, the insulin receptor is able to phosphorylate itself and turn itself ON!
That probably seems like an overly complicated way of turning on insulin signalling and telling the cell that its time to eat, but evolution is clever and biology is messy.