Enantioselective Binding of Designed Propranolol Analogues to the Cellobiohydrolase Cel7A

Sammanfattning: This thesis describes the affinity and enantiomeric selectivity of the cellobiohydrolase Cel7A (isolated from Hypocrea jercorina). The chiral recognition of the enzyme has been studied with respect to amino alcohols and new ligands have been designed, based on the structure of the common ?-blocker propranolol. Capillary electrophoresis have been used to determine the dissociation constants of the ligand binding at pH 5.0 and 7.0 to obtain information about the effects of the protonation state of the enzyme. A decrecased selectivity was shown at pH 7, despite an increase in affinity. The structural modifications in the ligands have also been related to the affinity and selectivity obtained. An expantion of the aromatic ring system of propranolol in combination with additional hydroxyl groups gave an increase in both affinity and selectivity. This resulted in an improved inhibitor of the enzyme. The energies involved in complexation, as in enthalpy and entropy, have been determined for a selected number of ligands and the entropy is the driving force of the selectivity, indicating the importnace of water displacement upon ligand binding. Also, new X-ray crystal structures have been obtained and structures of both enantiomers of the strongest binding ligand are now available. These structures have been used in computational calculations to study the ligand binding. The water molecules in the active site have been shown to have a much greater effect on the binding of the (R)-enantiomers, than for the (S)-enantiomers, which gives rise to the chiral selectivity of Cel7A.