Thermal Unfolding of Proteins
Probed at the Single Molecule Level Using Nanopores
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Abstract
The nanopore technique has great potential to discriminate
conformations of proteins. It is a very interesting system to mimic
and understand the process of translocation of biomacromolecules through
a cellular membrane. In particular, the unfolding and folding of proteins
before and after going through the nanopore are not well understood.
We study the thermal unfolding of a protein, probed by two protein
nanopores: aerolysin and α-hemolysin. At room temperature, the
native folded protein does not enter into the pore. When we increase
the temperature from 25 to 50 °C, the molecules unfold and the
event frequency of current blockade increases. A similar sigmoid function
fits the normalized event frequency evolution for both nanopores,
thus the unfolding curve does not depend on the structure and the
net charge of the nanopore. We performed also a circular dichroism
bulk experiment. We obtain the same melting temperature (around 45
°C) using the bulk and single molecule techniques