Cellobiose dehydrogenase on electrodes - an electrochemical biosensor for various analytes tunable by positive charges

Sammanfattning: Popular Abstract in English Enzymes are nanometer sized work units made up of proteins. Each of the thousands of the existing enzymes are produced by the cells of every living organism. They carry out certain functions as putting molecules together, e. g. connect sugar units to form cellulose for a growing tree or destroy molecules, e. g. degrade cellulose into its sugar units to eat and live on them. The latter is done by wood degrading fungi. One enzyme produced by such fungi and involved in the degradation process of wood is cellobiose dehydrogenase (CDH) – the main actor in this thesis, which is depicted on the cover of this thesis. The function of CDH is to substract electrons from cellobiose sugar units and deliver them to another enzyme, which makes highly reactive radicals with the help of those electrons and oxygen. Those oxygen radicals help to degrade the very stable wood. In this thesis CDH was put on electrode surfaces made of graphite and gold. Instead of delivering the electrons to the radical producing protein the electrons were delivered now to the electrode instead and the amount of gained electrons was counted by measuring the electrical current, which is nothing else than a flow of electrons. This is called a biosensor. Little amounts of sugar only give little currents and vice versa. So measuring the current one can calculate how much sugar is there. This is useful if one for example wants to know how much lactose is there in milk or how much sugar is there in blood. This is exactly how diabetric patients measure their blood sugar nowadays, only with a less fancy enzyme. In this thesis we found out that we can enhance the speed of how fast CDH can transport the electrons within the protein to the electrode by simply adding calcium ions, which are positively charged. That gave us more electric current so that we could measure sugar much more accurately. Also other positively charged compounds as long chained polymers gluing CDH to the electrode surface or to gold nanoparticles did the same effect. Our research showed that this effect depends on from which wood degrading fungus the enzyme was taken from, on the acidity of the measuring solution and on the calcium ion or polymer concentration. This is good to know because later sensors are supposed to only measure sugar units and should not be disturbed by calcium ions. In this thesis we also managed to open up another electron transfer pathway from deep inside the enzyme to the electrode. This helped us to get the electrons with less of an applied force, which is beneficial for the CDH biosensor because the less force we apply the smaller the chance of accidently substracting electrons from other molecules, which we don’t want to measure, like vitamin C. We also used a CDH biosensor to measure the release of lactose from ibuprofen tablets. Most tablets contain lactose as a filling material, only little of the tablet is the actual drug. So knowing how fast the lactose is released helps to understand how fast the tablet dissolves in the body. Another biosensor was made to measure ATP, the common energy currency in your body. But this was only possible using two other enzymes working together with CDH in an assembly line like manner on the electrode surface. The developed sensors are only prototypes – one cannot buy them. But in the near future CDH based biosensors are planned to be really used for measuring glucose in blood or lactose in dairy factories. And I look forward to participating in this.