offer many advantages and opportunities over lipid vesicles for all of these
applications (e.g., increased stability, tunable functionality, and permeability)
[87]. To date, many types of block copolypeptide amphiphiles that form stable
vesicular assemblies have been developed. The first of these utilized diethylene
glycol-modified lysine residues (i.e., K
P
) that impart both non-ionic water solubility
as well as ordered α-helical conformations to the hydrophilic polypeptide domains
[88]. Most other materials utilize highly charged polyelectrolyte segments to impart
both functionality and fluidity to the membranes. More recently, these copolypeptides
have included increasingly complex functionality to assist in cargo loading, vesicle
targeting, and vesicle disruption.
In 2004, Deming’s laboratory studied the roles of chain length and block
composition on the assembly of uncharged diblock copolypeptide amphiphiles of
the general structure poly(N ε -2-[2-(2-methoxyethoxy)ethoxy]acetyl-L-lysine)block-poly(L-leucine), or K
P
x L y [88]. These diblock copolypeptide amphiphiles
associate very strongly and essentially do not exist as single chains in aqueous
solution. This property, in most cases, results primarily in the formation of irregular
aggregates if the polymers are simply dispersed in deionized water. A protocol was
developed, using organic solvent (THF) and a denaturant (TFA) that allowed
annealing of these materials when water is added. Dialysis of the samples allows
one to obtain regular assemblies in pure water.
Using this procedure, a number of amphiphilic copolymers were studied in
which the hydrophilic domains were varied from 60 to 200 residues in average
length and the hydrophobic domains were varied from 10 to 75 residues in average
length [88]. All block copolypeptides were expected to adopt rod-like conformations due to the strong α-helix-forming tendencies of both the leucine and ethylene
glycol-modified lysine residues [27]. These rod-like conformations provided a
flat amphiphile interface upon association in water, thus directly tying polymer
conformation to supramolecular structure. Circular dichroism spectroscopy of the
copolymers in water confirmed that all samples were α-helical. Using differential
interference contrast (DIC) optical microscopy, TEM, laser scanning confocal
microscopy (LSCM), and DLS as initial methods for studying the assemblies,
some trends were identified [88]. When the hydrophobic poly(leucine) domains
were less than 20 residues in length, a significant fraction of oblong or irregular
micelles (ca. 100 nm diameter) formed, as observed by DLS and TEM. When the
size of the hydrophilic domain was 100 residues, unilamellar vesicles were
observed to form with a size range of approximately 2–15 μm diameter (Fig. 3).
When the hydrophilic block was increased to 150 residues, the vesicles were much
larger in size, approaching 50 μm in diameter. Finally, when the hydrophilic
segments were increased to 200 residues long, membrane curvature was hindered
such that the major structures formed were flat membrane sheets.
These block copolypeptides, where both hydrophilic and hydrophobic segments
were α-helical, gave rise to very stiff membranes, as suggested by the large vesicle
diameters and lack of fluidity in the sheets that were formed. Further investigation
revealed that these membranes were completely insensitive to osmotic stress, a
consequence of their impermeability to water, ions, or other small molecules
Synthesis and Self-Assembly of Well-Defined Block Copolypeptides via. . .
21
applications (e.g., increased stability, tunable functionality, and permeability)
[87]. To date, many types of block copolypeptide amphiphiles that form stable
vesicular assemblies have been developed. The first of these utilized diethylene
glycol-modified lysine residues (i.e., K
P
) that impart both non-ionic water solubility
as well as ordered α-helical conformations to the hydrophilic polypeptide domains
[88]. Most other materials utilize highly charged polyelectrolyte segments to impart
both functionality and fluidity to the membranes. More recently, these copolypeptides
have included increasingly complex functionality to assist in cargo loading, vesicle
targeting, and vesicle disruption.
In 2004, Deming’s laboratory studied the roles of chain length and block
composition on the assembly of uncharged diblock copolypeptide amphiphiles of
the general structure poly(N ε -2-[2-(2-methoxyethoxy)ethoxy]acetyl-L-lysine)block-poly(L-leucine), or K
P
x L y [88]. These diblock copolypeptide amphiphiles
associate very strongly and essentially do not exist as single chains in aqueous
solution. This property, in most cases, results primarily in the formation of irregular
aggregates if the polymers are simply dispersed in deionized water. A protocol was
developed, using organic solvent (THF) and a denaturant (TFA) that allowed
annealing of these materials when water is added. Dialysis of the samples allows
one to obtain regular assemblies in pure water.
Using this procedure, a number of amphiphilic copolymers were studied in
which the hydrophilic domains were varied from 60 to 200 residues in average
length and the hydrophobic domains were varied from 10 to 75 residues in average
length [88]. All block copolypeptides were expected to adopt rod-like conformations due to the strong α-helix-forming tendencies of both the leucine and ethylene
glycol-modified lysine residues [27]. These rod-like conformations provided a
flat amphiphile interface upon association in water, thus directly tying polymer
conformation to supramolecular structure. Circular dichroism spectroscopy of the
copolymers in water confirmed that all samples were α-helical. Using differential
interference contrast (DIC) optical microscopy, TEM, laser scanning confocal
microscopy (LSCM), and DLS as initial methods for studying the assemblies,
some trends were identified [88]. When the hydrophobic poly(leucine) domains
were less than 20 residues in length, a significant fraction of oblong or irregular
micelles (ca. 100 nm diameter) formed, as observed by DLS and TEM. When the
size of the hydrophilic domain was 100 residues, unilamellar vesicles were
observed to form with a size range of approximately 2–15 μm diameter (Fig. 3).
When the hydrophilic block was increased to 150 residues, the vesicles were much
larger in size, approaching 50 μm in diameter. Finally, when the hydrophilic
segments were increased to 200 residues long, membrane curvature was hindered
such that the major structures formed were flat membrane sheets.
These block copolypeptides, where both hydrophilic and hydrophobic segments
were α-helical, gave rise to very stiff membranes, as suggested by the large vesicle
diameters and lack of fluidity in the sheets that were formed. Further investigation
revealed that these membranes were completely insensitive to osmotic stress, a
consequence of their impermeability to water, ions, or other small molecules
Synthesis and Self-Assembly of Well-Defined Block Copolypeptides via. . .
21