Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 105, Issue 50, Pages 19720-19725Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.0808296105
Keywords
DNA sequencing; electroosmosis; nanopore; protein engineering; single-molecule detection
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Funding
- National Institutes of Health
- Medical Research Council
- Royal Society Wolfson Research Merit Award
- Medical Research Council [G0300122] Funding Source: researchfish
- MRC [G0300122] Funding Source: UKRI
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Both protein and solid-state nanopores are under intense investigation for the analysis of nucleic acids. A crucial advantage of protein nanopores is that site-directed mutagenesis permits precise tuning of their properties. Here, by augmenting the internal positive charge within the alpha-hemolysin pore and varying its distribution, we increase the frequency of translocation of a 92-nt single-stranded DNA through the pore at + 120 mV by approximate to 10-fold over the wild-type protein and dramatically lower the voltage threshold at which translocation occurs, e. g., by 50 mV for 1 event.s(-1).mu M-1. Further, events in which DNA enters the pore, but is not immediately translocated, are almost eliminated. These experiments provide a basis for improved nucleic acid analysis with protein nanopores, which might be translated to solid-state nanopores by using chemical surface modification.
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