Nanopore analysis of tethered peptides

• Published in: Journal of peptide science Vol. 16; no. 12; pp. 701 - 708
• Main Authors: Meng, Howard, Detillieux, Dielle, Baran, Christian, Krasniqi, Besnik, Christensen, Christopher, Madampage, Claudia, Stefureac, Radu I., Lee, Jeremy S.
• Format: Journal Article
• Language: English
• Published: Chichester, UK John Wiley & Sons, Ltd 01-12-2010
• Subjects: Amino acids; Benzene; Cysteine; Hemolysin Proteins - chemistry; Hemolysin Proteins - metabolism; Intercalation; Models, Molecular; Molecular Structure; nanopore; Nanostructures - chemistry; Nanotechnology; Peptides - analysis; Peptides - chemistry; Peptides - metabolism; Pores; tethered peptides; Translocation; α-hemolysin
• ISSN: 1075-2617; 1099-1387; 1099-1387
• DOI: 10.1002/psc.1289

Peptides of 12 amino acids were tethered via a terminal cysteine to mono‐, di‐, tri‐, and tetrabromomethyl‐substituted benzene to produce bundles of one to four peptide strands (CY12‐T1 to CY12‐T4, respectively). The interaction of the bundles with the α‐hemolysin pore was assessed by measuring the blockade currents (I) and times (T) at an applied potential of − 50, − 100, and − 150 mV. Three types of events could be distinguished: bumping events, with small I and short T where the molecule transiently interacts with the pore before diffusing away; translocation events, where the molecule threads through the pore with large I and the value of T decreases with increasing voltage; and intercalation events, where the molecule transiently enters the pore but does not translocate with large I and the value of T increases with increasing voltage. CY12‐T1 and CY12‐T2 gave only bumping and translocation events; CY12‐T3 and CY12‐T4 also gave intercalation events, some of which were of very long duration. The results suggest that three uncoiled peptide strands cannot simultaneously thread through the α‐hemolysin pore and that proteins must completely unfold in order to translocate. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd.
Threading the needle. Up to four short peptides were attached to a single benzene ring to produce peptide bundles and their ability to thread through the α‐hemolysin pore was assessed. Bundles of one or two strands readily translocated, whereas three‐ and four‐stranded bundles produced many bumping and intercalation events as well as permanent blockages. The results suggest that proteins must completely unfold in order to thread through the pore.