Dimerization of Proline Dehydrogenase from Thermus Thermophilus is Crucial for its Thermostability.

Thermus thermophilus proline dehydrogenase (TtProDH) catalyzes the first step in proline catabolism. The thermostable flavoenzyme consists of a distorted TIM-barrel and three N-terminal helices: αA, αB and αC. Using maltose-binding protein (MBP) fused constructs, we recently demonstrated that helix αC is crucial for TtProDH catalysis and for tetramerization through positioning of helix α8. Here, we report on the structural features that determine the thermostability of TtProDH. Selective disruption of two ion-pairs in the dimerization interface of several MBP-TtProDH variants resulted in the formation of monomers. The newly created monomers have improved catalytic properties but their melting temperatures are decreased by more than 20 °C. Sequence comparison suggested that one of the ion-pairs involved in dimerization is unique for ProDHs from Thermus species. In summary, intermolecular ion-pairs improve the thermostability of TtProDH and a trade-off is made between thermostability and catalytic activity. This article is protected by copyright. All rights reserved.