The Lipase B from Candida antarctica in Bicontinuous Microemulsions: a structural study

One of the limitations which biocatalysis faces nowadays arises from the fact that interesting hydrophobie substrates are insoluble in water whereas enzymes are soluble in water and often require an interface to be active. In this project we propose bicontinuous microemulsions as a reaction medium to successfully dissolve enzymes and non-natural substrates. Bicontinuous microemulsions consist of interpenetrating water and oil domains separated by a connected surfactant monolayer. In this study we introduced the lipase B from Candida antarctica (Cal B) into a bicontinuous microemulsion which consists of water - n-octane - nonionic surfactant (C10E5). The goals of this investigation were (1) to understand how Cal B influences the properties of bicontinuous microemulsions, i. e. the phase behaviour and the microstructure and (2) to investigate how the conformation of Cal B changes if it is introduced into a bicontinuous microemulsion. The respective phase diagrarns together with partitioning studies lead to the conclusion that around 80-9Owt.% ofthe initial amount of Cal B is located at the microemulsion's interface whereas the rest is located in the microemulsion's water domain. By knowing this, we compared the microstructure of lipase-free and lipase-containing microemulsions using PFG NMR and we found that the bicontinuity of the microemulsions is not influenced by the lipase. The conformational changes, which Cal B experiences in this reaction medium, were studied using circular dichroism spectroscopy. The results led to the conclusion that there is a continuous exchange between Cal B moleeules located at the interface and Cal B molecules located in the water domain of microemulsions.