Quantification of Aquaporin-Z reconstituted into vesicles for biomimetic membrane fabrication

Abstract

Aquaporin incorporated biomimetic membranes are anticipated to offer unprecedented desalination capabilities. However, the lack of accurate methods to quantify the reconstituted aquaporin presents a huge hurdle in investigating aquaporin performance and optimizing membrane fabrication. Herein, we present three quantification methods to determine the Aquaporin-Z reconstituted into E. coli lipid vesicles: 1) nanogold labeling with transmission electron microscopy (TEM) visualization, 2) nickel labeling with inductively coupled plasma-mass spectrometry (ICP-MS) and 3) gel electrophoresis. The TEM method serves as a quick way to determine if aquaporin has been reconstituted, but is not quantitative. The numerical results from quantitative methods, ICP-MS and gel electrophoresis, correlate closely, showing that 60 ± 20% vs 66 ± 4% of Aquaporin-Z added is successfully reconstituted into vesicles respectively. These methods allow more accurate determination of Aquaporin-Z reconstituted and loss during reconstitution, with relatively commonly available equipment and without complex sample handling, or lengthy data analysis. These would allow them to be widely applicable to scientific studies of protein function in the biomimetic environment and engineering studies on biomimetic membrane fabrication.