KT 323 houses the biochemistry preparation room and a large part of the equipment for protein expression and isolation.

To produce proteins, e.g. enzymes that catalyse a particular reaction, they can be extracted from a natural source (microorganisms/plants/tissue). Another possibility is a process known as heterologous expression. In order to achieve this, the gene for the protein is transferred to another organism and the protein is produced there. Micro organisms such as E. coli or unicellular fungi are particularly suitable for this purpose.  

After cultivation, the cells of the protein-producing bacteria or yeasts are destroyed and the proteins are purified using chromatographic methods.

Protein purification by low-pressure LC

Nowadays, chromatographic methods are mainly used for protein purification. In the Biochemistry laboratory, two low-pressure liquid chromatography systems are used. Here, important separations such as affinity chromatography and ion exchange chromatography can be carried out using a minimal amount of equipment. For the analysis of sensitive samples, all systems are operated at 4°C.

Protein extracts, e.g. from cultures that express heterologous proteins, are applied to the columns required in each case. The desired proteins bind; impurities pass through the column.  Finally, the desired proteins are dissolved (eluted) from the column in enriched form. The eluted proteins are automatically collected in fractions. Within these protein fractions, the content of the desired protein is then determined, e.g. by activity tests, SDS-Page, or western blot.

SDS-polyacrylamide gel electrophoresis

To investigate the composition of protein extracts or fractions from protein chromatography, samples thereof are applied to a polyacrylamide gel and separated by electrophoresis in the presence of the strong detergent SDS. Due to the binding of the SDS, all proteins are then strongly negatively charged and migrate to the positive pole in the electric field. The polyacrylamide gel causes small proteins to migrate faster than large proteins. By comparing them with proteins of known size, the molecular weight of the proteins contained can therefore be determined approximately.


In the Biochemistry laboratory, high-pressure liquid chromatography (HPLC) is especially used to examine samples with small molecules on an analytical scale. We mainly use reversed phase (RP) chromatography on C18 column material. The more hydrophobic a substance is, the better it binds to this hydrophobic material. By continuously increasing the hydrophobic portion in the solvent (usually acetonitrile in water), the elution power of the liquid phase is then increased and the more hydrophobic compounds are gradually released from the column. Eluted substances are quantified by measuring the absorption in a diode array detector.

If necessary, HPLC can also be operated on a preparatory scale after replacing the pump heads and connecting a fraction collector.