Urea facilitates the translocation of single-stranded DNA and RNA through the alpha-hemolysin nanopore.
The staphylococcal alpha-hemolysin (alphaHL) 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 alphaHL 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 alphaHL pore in the presence of 4 M urea, a concentration that denatures the secondary structure of the polynucleotides. The alphaHL pore is functional even in 7 M urea, and therefore it is easily stable enough for analyses of challenging DNA and RNA species.
Japrung
Deanpen
D
Department of Chemistry, University of Oxford, Oxford, United Kingdom.
Henricus
Marsiyana
M
Li
Qiuhong
Q
Maglia
Giovanni
G
Bayley
Hagan
H
D001427:Bacterial Toxins / Q000737:chemistry* / Q000378:metabolism*; D001483:Base Sequence; D001692:Biological Transport / Q000187:drug effects; D002942:Circular Dichroism; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004553:Electric Conductivity; D004591:Electrophoresis, Polyacrylamide Gel; D006460:Hemolysin Proteins / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D049329:Nanostructures; D016062:Porosity; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D012313:RNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012996:Solutions; D013050:Spectrometry, Fluorescence; D013211:Staphylococcus aureus; D014508:Urea / Q000494:pharmacology*
D001427:Bacterial Toxins; D004277:DNA, Single-Stranded; D006460:Hemolysin Proteins; D012996:Solutions; C021091:staphylococcal alpha-toxin; D012313:RNA; D014508:Urea