Biomedical Engineering Reference
In-Depth Information
6.1
Introduction
The combination of variable polypeptide sequence and nearly arbitrary chain length
results in an astronomical number of possible proteins. The resulting variety of
structures and functions of proteins are so complicated that in some ways the
diversity of life on Earth can be viewed as a consequence thereof. Eukaryotic life
forms must transport functional biopolymers across membranes to survive. This
fact motivates study of protein translocation through nanometer-scale pores. Such
understanding is of fundamental importance not only for basic science but also in
biotechnological applications that seek to mimic the selectivity and sensitivity of
biological translocation [
1
,
2
].
The physiochemical properties of the two most important classes of biopoly-
mers, polynucleic acids and polypeptides, are substantially different. Therefore the
cellular transport machinery is different depending on the nature of the biopolymer
being transported. Proteins in particular present special challenges for cells, as they
must be transported in a way that is compatible with attainment of a correctly folded
three-dimensional native structure. The experimental and theoretical approaches to
both natural and artificial ion channel translocation must accommodate these
differences.
In this chapter we summarize the recent development of single nanopore
measurements and theory to enable measurement of the physical properties of
proteins at the single molecule level. These physical properties include: the protein
size or volume, electrical charge, and conformational states.
6.1.1 Polypeptides Measured by Protein Pores
Protein channels or protein pores such as
-hemolysin have well defined structure
and dimensions. However, due to their small fixed diameter, only polypeptides or
denatured proteins are able to pass through the pores. Several research groups have
studied polypeptide and protein pore interactions and the results have been
presented in many publications. Starting in the 1990s, experiments began revealing
that some peptide chains could reside inside in the lumen of protein pores or
channels [
3
]. Later, studies in Lee's lab demonstrated that resistive pulse signals
from a single
a
-hemolysin pore could differentiate between single, double, and
collagen-like triple helices and pulses from
a
-hemolysin or aerolysin pores could
reveal differences between wild type and mutant Histidine containing protein [
4
,
5
].
The Auvray and Pelta research groups have studied the interaction of dextran
sulfate and maltose binding protein with
a
-hemolysin pores [
6
,
7
]. The Movileanu
and Bayley laboratories have used wild type and mutated
a
-hemolysin pores to
examine the effect of electrostatics on the interaction between peptide sequences
and the
a
a
-hemolysin pore [
8
,
9
].
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