Monday, October 17, 2011: 8:30 AM
208 D (Minneapolis Convention Center)
In this work, a new approach of incorporating transmembrane protein AquaporinZ (AqpZ) into the lipid bilayer has been developed with the aid of Langmuir-Blodgett technique. Aquaporin is a big family of transmembrane proteins that selectively conduct water or small solutes in or out of the biological membrane driven by osmotic pressures. The hydrophobic property of AqpZ makes the incorporation of membrane proteins with the LB method a challenging and intriguing work. The incorporation in this study is achieved by combining a pure binary-lipid monolayer with an AqpZ-associated binary-lipid monolayer and subsequent refolding of AqpZ in the bilayer. The binary-lipid monolayer is composed of (1) gel-phase lipids that prevent detergent dissolution and (2) nickel-chelating lipids that attach the AqpZ from the subphase. To gain better film integrity and enhance protein adsorption on the monolayer, detergent in the subphase is removed by BioBeads. It is shown that the removal rate can be determined by BioBeads amount and subphase circulation. Furthermore, the bilayer formed with reconstituted AqpZ is imaged by AFM and the incorporation mechanism is explained in a three-step process. The new approach of AqpZ reconstitution revealed by this work could potentially be applied in biomimetic membranes for water purification and biosensor applications.
Acknowledgement This work was financially supported by the National Research Foundation of Singapore, Environment and Water Industry Programme Office (EWI) project 0804-IRIS-01 (NUS grant number: R-279-000-293-272).