Sensing Single Protein Molecules with Solid-State Nanopores - Nanopores: Sensing and Fundamental Biological Interactions - page 148

The current blockage signal from a nanopore measurement contains the
information on the protein's shape or folding state and on the sequence of the
amino-acid volume and charge of the local segment of the polypeptide chain in
the pore if the protein molecule is unfolded. The research on using solid-state
nanopores to measure the shape of single protein molecules at different folding
states and to probe its charge sequence when it is unfolded in salt solution is still at a
very early stage.
Equations (6.1), (6.3) and (6.4) were developed using several assumptions
including: (1) a uniform distribution of charge on the protein, (2). the protein is a
small hard sphere, (3) the protein does not interact with the pore walls, and (4) there
is only one ionization state of the protein. These assumptions limit the general
applicability of these equations. Future work must evaluate the need for corrections
for these assumptions to enable a better description of protein translocation.
Complicated assemblies of proteins such as laminin produce complex signals
that cannot be uniquely interpreted. For example, the recorded current trace
(Fig. 6.1c ) for Laminin protein shows the complexity of the signal measured for a
partially denatured protein. At the present time, we are still developing analysis
routines to process these more complicated data sets. In this chapter we only
attempted to discuss the current blockage signal produced by proteins in two
simplest conformations: in their native state and in their completely unfolded state.
Future developments of nanopore techniques for sensing proteins should be
focused on improvement of signal-to-noise, understanding of protein translocation
signal and protein structure, dynamic adjustment of DC bias potential, control of
nanopore surface chemistry, in situ characterization of nanopore geometry and
electrical response, analysis improvements to provide richer translocation data,
and incorporation of other single molecule methods into the nanopore experiments.

Acknowledgments We thank Professor J. Golovchenko and Harvard nanopore group for nanopore
fabrication, Ryan Rollings, Edward W. Graef Jr., Denis F. Tita, and Errol Porter for nanopore
fabrication and characterization. We acknowledge the funding support provided by NHGRI/NIH
R21HG003290, NHGRI/NIH R21HG00477, NSF/MRSEC 080054, ABI-111/710, and NIH
R01GM071684 to DST.

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