Title: Detection of Two Isomeric Binding Configurations in a Protein-Aptamer Complex with a Biological Nanopore
Authors: Van Meervelt, Veerle
Soskine, Misha
Maglia, Giovanni # ×
Issue Date: Dec-2014
Publisher: American Chemical Society
Series Title: ACS Nano vol:8 issue:12 pages:12826-35
Abstract: Protein-DNA interactions play critical roles in biological systems, and they often involve complex mechanisms and dynamics that are not easily measured by ensemble experiments. Recently, we showed that folded proteins can be internalised inside ClyA nanopores and studied by ionic current recordings at the single-molecule level. Here, we use ClyA nanopores to sample the interaction between the G-quadruplex fold of the thrombin binding aptamer (TBA) and human thrombin (HT). Surprisingly, the internalisation of the HT:TBA complex inside the nanopore induced two types of current blockades with distinguished residual current and lifetime. Using single nucleobase substitutions to TBA we showed that these two types of blockades originate from TBA binding to thrombin with two isomeric orientations. Voltage dependencies and the use of ClyA nanopores with two different diameters allowed assessing the effect of the applied potential and confinement, and revealed that the two binding configurations of TBA to HT display different lifetimes. These results show that the ClyA nanopores can be used to probe conformational heterogeneity in protein:DNA interactions.
ISSN: 1936-0851
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Biochemistry, Molecular and Structural Biology Section
× corresponding author
# (joint) last author

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