Arrayed lipid bilayer chambers allow single-molecule analysis of membrane transporter activity

• Published in: Nature communications 2014-07, Vol.5 (1), p.4519, Article 4519
• Main Authors: Watanabe, Rikiya, Soga, Naoki, Fujita, Daishi, Tabata, Kazuhito V., Yamauchi, Lisa, Hyeon Kim, Soo, Asanuma, Daisuke, Kamiya, Mako, Urano, Yasuteru, Suga, Hiroaki, Noji, Hiroyuki
• Format: Article
• Language: English
• Subjects: 631/57/2265; 631/57/2270; 631/80; 631/92/577; 9/10; Bacterial Toxins - metabolism; Biological Transport, Active; Fluorescent Dyes - metabolism; Hemolysin Proteins - metabolism; Humanities and Social Sciences; Hydrazines - metabolism; Lipid Bilayers - metabolism; Membrane Transport Proteins - analysis; Membrane Transport Proteins - metabolism; Microtechnology; Molecular Biology - instrumentation; Molecular Biology - methods; multidisciplinary; Nanotechnology - instrumentation; Nanotechnology - methods; Photobleaching; Proton-Translocating ATPases - metabolism; Science; Science (multidisciplinary)
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/ncomms5519

Nano- to micron-size reaction chamber arrays (femtolitre chamber arrays) have facilitated the development of sensitive and quantitative biological assays, such as single-molecule enzymatic assays, digital PCR and digital ELISA. However, the versatility of femtolitre chamber arrays is limited to reactions that occur in aqueous solutions. Here we report an arrayed lipid bilayer chamber system (ALBiC) that contains sub-million femtolitre chambers, each sealed with a stable 4-μm-diameter lipid bilayer membrane. When reconstituted with a limiting amount of the membrane transporter proteins α-hemolysin or F 0 F 1 -ATP synthase, the chambers within the ALBiC exhibit stochastic and quantized transporting activities. This demonstrates that the single-molecule analysis of passive and active membrane transport is achievable with the ALBiC system. This new platform broadens the versatility of femtolitre chamber arrays and paves the way for novel applications aimed at furthering our mechanistic understanding of membrane proteins’ function. The development of small volume chamber arrays has greatly facilitated high throughput biological assays of soluble proteins. Here, Watanabe et al. adapt this approach to develop an arrayed lipid bilayer chamber system for single molecule level measurements of membrane transporter activity.