Prenyltransferases catalyze the formation of C-C-bonds between an electron-rich acceptor and allylic isoprenoid diphosphates as electrophiles. In contrast, terpene-cyclases are mainly responsible for the cyclization of iosprenoid compounds to a huge variety of cyclic mono-, di-, tri- and oligoterpenes. Nevertheless, both enzyme families belong to the main group of prenylating enzymes, which products are ultimately converted into over 60000 naturally occurring isoprenoid containing compounds - essential in all organisms. In E. coli, for instance, the isoprenoids ubiquinone and bactoprenol are required for respiration and cell wall biosynthesis, respectively. Up to now, for the investigated enzymes there is no X-ray structure known. Only limited similarities or in very few cases homologous proteins could be found. Molecular modeling in combination with site directed mutagenesis should help to explain the substrate specificity and to explore the evolutionary origin of both classes of enzymes. Based on these modeling analyses, amino acids identified as important to the catalysis mechanism were selectively replaced to obtain new mutants. All mutants were tested for their ability to form the expected products and then compared to the wildtype-enzymes. The results in all cases confirm the involvement of the mutated amino acids in the catalysis mechanism and were used as basis for comprehensive quantum mechanical investigations (docking, QM/MM). Finally all findings allow a deeper insight into the catalysis mechanism of prenylating enzymes and might help to design specific ligands (e.g. inhibitors) for this important class of biocatalysts.