The rapid quantification of the enantiomers of small chiral molecules is very important, notably in pharmacology. Here, we show that the enantiomers of drug molecules can be distinguished by stochastic sensing, a single-molecule detection technique. The sensing element is an engineered alpha-hemolysin protein pore, fitted with a beta-cyclodextrin adapter. By using the approach, the enantiomeric composition of samples of ibuprofen and thalidomide can be determined in less than 1 s.
Here we report a rapid, label-free method for monitoring peptide cleavage. Monitoring peptide translocation through an engineered ion channel in the absence and the presence of an enzyme allowed quantitative chemical kinetics information on enzymatic processes to be obtained. In addition to its potential application in disease diagnostics and drug discovery, this peptide/protein cleavage approach is envisioned for further development as a novel rapid, label-free protein sequencing technique.
Engineered versions of the transmembrane protein pore alpha-hemolysin (alphaHL) can be used as stochastic sensing elements for the identification and quantification of a wide variety of analytes at the single-molecule level. Until now, nitroaromatic analytes have eluded detection by this approach. We now report that binding sites for nitroaromatics can be built within the lumen of the alphaHL pore from simple rings of seven aromatic amino acid side chains (Phe, Tyr or Trp). By monitoring the ionic current that passes through a single pore at a fixed applied potential, various nitroaromatics can be distinguished from TNT on the basis of the amplitude and duration of individual current-blocking events. Rings of less than seven aromatics bind the analytes more weakly; this suggests that direct aromatic-aromatic interactions are involved. The engineered pores should be useful for the detection of explosives and, in combination with computational approaches and structural analysis, they could further our understanding of noncovalent interactions between aromatic molecules.
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