Enzymology is the purification and structural/functional characterization of enzymes. Techniques are sonication and the french press, to obtain cell free extract, ultra-centrifugation to separate membrane from soluble proteins, initial separation by ammoniumsulfate precipitation, FPLC (anion/cation exchange, hydrophobic interaction, gel filtration), HPLC, and several concentration methods. Proteins are assayed by 1D or 2D gel electrophoresis, combined with colorimetric protein concentration determinations. Functional characterization of proteins is carried out in activity assays, with UV/VIS spectroscopy (using a photo-diode array spectrophotometer). For further characterization (such as EPR) we cooperate with Simon de Vries, at the TU Delft. The MALDI-TOF is a convenient way to go from the protein to the genetic level: via peptide-mapping we can obtain the gene sequences of purified proteins.
