Stochastic sensing of organic analytes by a pore-forming protein containing a molecular adapter

The detection of organic molecules is important in many areas, including me dicine, environmental monitoring and defence(1-5) Stochastic sensing is an approach that relies on the observation of individual binding events betwee n analyte molecules and a single receptor(6). Engineered transmembrane prot ein pores are promising sensor elements for stochastic detection(6), and in their simplest manifestation they produce a fluctuating binary ('on/off') response in the transmembrane electrical current. The frequency of occurren ce of the fluctuations reveals the concentration of the analyte, and its id entity can be deduced from the characteristic magnitude and/or duration of the fluctuations. Genetically engineered versions of the bacterial pare-for ming protein alpha-haemolysin have been used to identify and quantify dival ent metal ions in solution(6). But it is not immediately obvious how versat ile binding sites for organic ligands might be obtained by engineering of t he pore structure. Here we show that stochastic sensing of organic molecule s can be procured from alpha-haemolysin by equipping the channel with an in ternal, non-covalently bound molecular 'adapter' whim mediates channel bloc king by the analyte. We use cyclodextrins as the adapters because these fit comfortably inside the pore and present a hydrophobic cavity suitable for binding a variety of organic analytes. Moreover, at single sensing element of this sort can be used to analyse a mixture of organic molecules with dif ferent binding characteristics. We envisage the use of other adapters, so t hat the pore could be 'programmed' for a rang of sensing functions.