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Stochastic Detection of Motor Protein–RNA Complexes by Single‐Channel Current Recording
Author(s) -
Astier Yann,
Kainov Denis E.,
Bayley Hagan,
Tuma Roman,
Howorka Stefan
Publication year - 2007
Publication title -
chemphyschem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.016
H-Index - 140
eISSN - 1439-7641
pISSN - 1439-4235
DOI - 10.1002/cphc.200700179
Subject(s) - rna , nucleic acid , chemistry , molecular motor , biophysics , dissociation (chemistry) , dna , atpase , biochemistry , nanotechnology , enzyme , biology , materials science , gene
A label‐ and immobilization‐free approach to detecting the reversible formation of complexes between nucleic acids and proteins at the single‐molecule level is described. The voltage‐driven translocation of individual oligoribonucleotides through a nanoscale protein pore is observed by single‐channel current recordings. The oligoribonucleotide 5′‐C 25 A 25 ‐3′ gives rise to current blockades with an average duration of approximately 0.5 ms. In the presence of the RNA‐binding ATPase P4, a viral packaging motor from bacteriophage ϕ8, longer events of tens to hundreds of milliseconds are observed. Upon addition of ATP the long events disappear, indicating the dissociation of the P4⋅RNA complex. The frequency of events also depends on the concentration of P4 and the length of the oligoribonucleotide, thereby confirming the specificity of the P4⋅RNA events. This study shows that single‐channel current recordings can be used to monitor RNA–protein complex formation, thus opening up a new means to examine the motor activity of RNA‐ or DNA‐processing enzymes.