Author:

Keywords:

nanometer-scale pore, polynucleotide molecules, secondary structure, hairpin molecules, aqueous solutions, protein pore, acid, discrimination, viscosity, database, dna sequencing, nanopore, protein engineering, single-nucleobase discrimination, protein-rna interactions, dna-binding domain, biological nanopore, single, base, molecules, strand, Science & Technology, Life Sciences & Biomedicine, Biophysics, AQUEOUS-SOLUTIONS, PROTEIN PORE, MOLECULES, DISCRIMINATION, VISCOSITY, SECONDARY, ACID, CONDUCTIVITY, DENSITY, GLYCINE, Bacterial Toxins, Base Sequence, Biological Transport, Circular Dichroism, DNA, Single-Stranded, Electric Conductivity, Electrophoresis, Polyacrylamide Gel, Hemolysin Proteins, Models, Molecular, Molecular Sequence Data, Nanostructures, Porosity, Protein Multimerization, Protein Structure, Quaternary, RNA, Solutions, Spectrometry, Fluorescence, Staphylococcus aureus, Urea, 02 Physical Sciences, 03 Chemical Sciences, 06 Biological Sciences, 31 Biological sciences, 34 Chemical sciences, 51 Physical sciences

Abstract:

The staphylococcal a-hemolysin (alpha HL) protein nanopore is under investigation as a fast, cheap detector for nucleic acid analysis and sequencing. Although discrimination of all four bases of DNA by the alpha HL pore has been demonstrated, analysis of single-stranded DNAs and RNAs containing secondary structure mediated by basepairing is prevented because these nucleic acids cannot be translocated through the pore. Here, we show that a structured 95-nucleotide single-stranded DNA and its RNA equivalent are translocated through the alpha HL pore in the presence of 4 M urea, a concentration that denatures the secondary structure of the polynucleotides. The alpha HL pore is functional even in 7 M urea, and therefore it is easily stable enough for analyses of challenging DNA and RNA species.